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<SOH> BRIEF DESCRIPTION OF THE DRAWINGS <EOH>FIG. 1 is a diagram showing the inhibitory effect of Calea extract on adipocyte differentiation induction. The ordinate shows 14 C—CH 3 COOH incorporation (cpm). “−Ins.”, “+Ins.”, and “Ins.+” on the abscissa indicate no addition of insulin, addition of insulin, and addition of both insulin and Calea extract to the sample, respectively. FIG. 2 is a diagram showing the inhibitory effect of Calea extract on adipocyte differentiation induction. The ordinate shows 14 C-2-deoxyglucose incorporation (cpm). “−Ins.”, “+Ins.”, and “Ins.+” on the abscissa indicate no addition of insulin, addition of insulin, and addition of both insulin and Calea extract to the sample, respectively. FIG. 3 is a diagram showing the isolation process of compounds (sesquiterpenoid derivatives) having an inhibitory effect on adipocyte differentiation induction. FIG. 4 is a diagram showing the inhibitory effect of sesquiterpenoid derivatives on adipocyte differentiation induction. The ordinate shows 14 C—CH 3 COOH incorporation (cpm/mg protein) “−Ins.”, and “+Ins.” on the abscissa indicate no addition of insulin, and addition of insulin to the sample, respectively. 1, 3, 4, 6, 7, 11, and 12 on the abscissa indicate samples to which compounds 1, 3, 4, 6, 7, 11, and 12, respectively, have been added in combination with insulin. detailed-description description="Detailed Description" end="lead"? |
Printing cartridge with capacitive sensor identification |
A printing cartridge (1230) includes a housing (1231). An actuating formation (1240) is positioned on the housing and is capable of actuating a number of capacitive sensors (1238) in an array of such sensors. The actuating formation is configured to represent data relating to at least one of: a serial number of the cartridge, a media and a media colorant, so that the capacitive sensors, when actuated, together generate a signal carrying such data |
1. A printing cartridge that comprises a housing; and an actuating formation that is positioned on the housing and is capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to at least one of: a serial number of the cartridge, a media and a media colorant, so that the capacitive sensors, when actuated, together generate a signal carrying such data. 2. A method of determining a media colorant of a printing cartridge, the method comprising the step of actuating a number of capacitive sensors within an array of such sensors with an actuating formation positioned on the printing cartridge, the actuating formation representing data relating to a media colorant in the cartridge, so that the capacitive sensors, when actuated, generate a signal carrying data relating to the media colorant. 3. A printing cartridge that comprises a housing; a media colorant supply arrangement positioned within the housing and containing a supply of media colorant; and an actuating formation positioned on the housing and being capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to the media colorant so that a signal generated by the capacitive sensors, when actuated, carries said data relating to the media colorant. 4. A printing cartridge as claimed in claim 3, wherein the data represented by the actuating formation relates to at least one of: a serial number identifying the media colorant, a type of the media colorant, a viscosity of the media colorant, a surface tension of the media colorant, optical characteristics of the media colorant and an optimal ink drop volume corresponding to a type of media. 5. A printing cartridge as claimed in claim 3, wherein a conductive material defines the actuating formation so that the actuating formation and a capacitive plate of each of said number of capacitive sensors define a capacitor. 6. A printing cartridge as claimed in claim 5, in which the actuating formation is defined by a plurality of projections that extend from the housing in an array which represents the data, each projection corresponding with a capacitive plate of each capacitive sensor of said number of capacitive sensors. 7. A printing cartridge as claimed in claim 3, in which the actuating formation is the product of an injection micromolding process. 8. A method of determining media of a printing cartridge, the method comprising the step of actuating a number of capacitive sensors in an array of such sensors with an actuating formation positioned on the printing cartridge, the actuating formation representing data relating to the media in the cartridge, so that the capacitive sensors, when actuated, generate a signal carrying data relating to the media. 9. A printing cartridge that comprises a housing; a media supply arrangement positioned within the housing and containing a supply of media; and an actuating formation positioned on the housing and being capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to the media so that a signal generated by the capacitive sensors, when actuated, carries said data relating to the media. 10. A printing cartridge as claimed in claim 9, wherein the data represented by the actuating formation relates to at least one of: a serial number identifying the media, a type of the media and a length of the media. 11. A printing cartridge as claimed in claim 9, wherein a conductive material defines the actuating formation so that the actuating formation and a capacitive plate of each of said number of capacitive sensors define a capacitor. 12. A printing cartridge as claimed in claim 11, in which the actuating formation is defined by a plurality of projections that extend from the housing in an array which represents the data, each projection corresponding with a capacitive plate of each capacitive sensor of said number of capacitive sensors. 13. A printing cartridge as claimed in claim 9, in which the actuating formation is the product of an injection micromolding process. 14. A method of determining media and media colorant of a printing cartridge, the method comprising the step of actuating a number of capacitive sensors in an array of such sensors with an actuating formation positioned on the printing cartridge, the actuating formation representing data relating to the media and the media colorant in the cartridge, so that the capacitive sensors, when actuated, generate a signal carrying data relating to the media and the media colorant. 15. A printing cartridge that comprises a housing; media and media colorant supply arrangements positioned within the housing and containing a supply of media and a supply of media colorant, respectively; and an actuating formation positioned on the housing and being capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to the media and the media colorant so that a signal generated by the capacitive sensors, when actuated, carries said data relating to the media and the media colorant. 16. A printing cartridge as claimed in claim 15, wherein the data represented by the actuating formation relates to at least one of: a serial number identifying the media, a serial number identifying the media colorant, a length of the media, a type of the media, a viscosity of the media colorant, a surface tension of the media colorant, optical characteristics of the media colorant and an optimal ink drop volume of the media colorant corresponding to the type of media. 17. A printing cartridge as claimed in claim 15, wherein a conductive material defines the actuating formation so that the actuating formation and a capacitive plate of each of said number of capacitive sensors define a capacitor. 18. A printing cartridge as claimed in claim 17, in which the actuating formation is defined by a plurality of projections that extend from the housing in an array which represents the data, each projection corresponding with a capacitive plate of each capacitive sensor of said number of capacitive sensors. 19. A printing cartridge as claimed in claim 15, in which the actuating formation is the product of an injection micromolding process. 20. A printing device that comprises a body, a printing cartridge being engageable with the body, the printing cartridge having a housing, a media colorant supply arrangement positioned within the housing and containing a supply of media colorant, an actuator formation being positioned on the housing and representing data relating to the media colorant; a processor positioned in the body to control operation of a media colorant feed mechanism and a printing mechanism; and an array of capacitive sensors positioned in the body and being configured so that predetermined combinations of capacitive sensors in the sensor array, when actuated, generate signals carrying data related to the media colorant, such predetermined combinations of capacitive sensors in the sensor array being actuable by the actuator formation positioned on the housing of the printing cartridge when the printing cartridge is engaged with the body so that the array of capacitive sensors generates a signal carrying said data relating to the media colorant of the printing cartridge. 21. A printing device as claimed in claim 20, in which the array of capacitive sensors is the product of an integrated circuit fabrication process. 22. A printing device as claimed in claim 21, in which the array of capacitive sensors is in the form of a ceramic metal oxide semiconductor (CMOS) device. 23. A printing device as claimed in claim 21, in which the array of capacitive sensors includes a substrate having dielectric properties, the substrate defining a contact surface against which the actuating formation bears, with each capacitive sensor including a capacitor plate positioned in the substrate, and spaced from the contact surface, so that, when the actuating formation bears against the contact surface, the capacitor plate and the actuating formation defines a capacitor. 24. A printing device as claimed in claim 23, in which the capacitor plates are positioned so that capacitor plates of predetermined combinations of capacitor plates correspond with projections of the actuating formation, to define capacitors having a capacitance that represents the data relating to the media colorant 25. A printing device as claimed in claim 23, in which the array of capacitive sensors incorporates circuitry to determine said capacitance. 26. A printing device that comprises a body, a printing cartridge being engageable with the body, the printing cartridge having a housing, a media supply arrangement positioned within the housing and containing a supply of media, an actuating formation being positioned on the housing and representing data relating to the media; a processor positioned in the body to control operation of a media feed mechanism and a printing mechanism; and an array of capacitive sensors positioned in the body and being configured so that predetermined combinations of capacitive sensors in the sensor array, when actuated, generate signals carrying data related to the media, such predetermined combinations of capacitive sensors in the sensor array being actuable by the actuator formation positioned on the housing of the printing cartridge when the printing cartridge is engaged with the body so that the array of capacitive sensors generates a signal carrying said data relating to the media of the printing cartridge. 27. A printing device as claimed in claim 26, in which the array of capacitive sensors is the product of an integrated circuit fabrication process. 28. A printing device as claimed in claim 27, in which the array of capacitive sensors is in the form of a ceramic metal oxide semiconductor (CMOS) device. 29. A printing device as claimed in claim 27, in which the array of capacitive sensors includes a substrate having dielectric properties, the substrate defining a contact surface against which the actuating formation bears, with each capacitive sensor including a capacitor plate positioned in the substrate, and spaced from the contact surface, so that, when the actuating formation bears against the contact surface, the capacitor plate and the actuating formation defines a capacitor. 30. A printing device as claimed in claim 23, in which the capacitor plates are positioned so that capacitor plates of predetermined combinations of capacitor plates correspond with projections of the actuating formation, to define capacitors having a capacitance that represents the data relating to the media. 31. A printing device as claimed in claim 23, in which the array of capacitive sensors incorporates circuitry to determine said capacitance. 32. A printing device that comprises a body, a printing cartridge being engageable with the body, the printing cartridge having a housing, media colorant and media supply arrangements positioned within the housing and containing a supply of media and media colorant, an actuating formation being positioned on the housing and representing data relating to the media colorant and the media; a processor positioned in the body to control operation of media colorant and media feed mechanisms and a printing mechanism; and an array of capacitive sensors positioned in the body and being configured so that predetermined combinations of capacitive sensors in the sensor array, when actuated, generate signals carrying data related to the media and the media colorant, such predetermined combinations of capacitive sensors in the sensor array being actuable by the actuator formation positioned on the housing of the printing cartridge when the printing cartridge is engaged with the body so that the array of capacitive sensors generates a signal carrying said data relating to the media and the media colorant of the printing cartridge. 31. A printing device as claimed in claim 30, in which the array of capacitive sensors is the product of an integrated circuit fabrication process. 32. A printing device as claimed in claim 31, in which the array of capacitive sensors is in the form of a ceramic metal oxide semiconductor (CMOS) device. 33. A printing device as claimed in claim 31, in which the array of capacitive sensors includes a substrate having dielectric properties, the substrate defining a contact surface against which the actuating formation bears, with each capacitive sensor including a capacitor plate positioned in the substrate, and spaced from the contact surface, so that, when the actuating formation bears against the contact surface, the capacitor plate and the actuating formation defines a capacitor. |
<SOH> BACKGROUND OF THE INVENTION <EOH>Recently, digital printing technology has been proposed as a suitable replacement for traditional camera and photographic film techniques. The traditional film and photographic techniques rely upon a film roll having a number of pre-formatted negatives which are drawn past a lensing system and onto which is imaged a negative of a image taken by the lensing system. Upon the completion of a film roll, the film is rewound into its container and forwarded to a processing shop for processing and development of the negatives so as to produce a corresponding positive set of photos. Unfortunately, such a system has a number of significant drawbacks. Firstly, the chemicals utilized are obviously very sensitive to light and any light impinging upon the film roll will lead to exposure of the film. They are therefore required to operate in a light sensitive environment where the light imaging is totally controlled. This results in onerous engineering requirements leading to increased expense. Further, film processing techniques require the utilizing of a “negative” and its subsequent processing onto a “positive” film paper through the utilization of processing chemicals and complex silver halide processing etc. This is generally unduly cumbersome, complex and expensive. Further, such a system through its popularity has lead to the standardization on certain size film formats and generally minimal flexibility is possible with the aforementioned techniques. Recently, all digital cameras have been introduced. These camera devices normally utilize a charge coupled device (CCD) or other form of photosensor connected to a processing chip which in turn is connected to and controls a media storage device which can take the form of a detachable magnetic card. In this type of device, the image is captured by the CCD and stored on the magnetic storage device. At some later time, the image or images which have been captured are down loaded to a computer device and printed out for viewing. The digital camera has the disadvantage that access to images is non-immediate and the further post processing step of loading onto a computer system is required, the further post processing often being a hindrance to ready and expedient use. Therefore, there remains a general need for an improved form of camera picture image production apparatus which is convenient, simple and effective in operation. Further, there also remains a need for a simple form of portable, immediate print media on which images can be effectively reproduced. In the parent application, there is disclosed the use of an authentication chip to provide information in connection with the print media and the media colorant that is supplied with the cartridge. The Applicant has identified that it would be highly desirable to provide a means whereby information concerning one or both of the media and the media colorant could be supplied together with the cartridge. The reason for this is that such information could be used, in a suitable form, by a processor of such a device to enhance operation of a printing mechanism. It will be appreciated that printing mechanisms need to operate differently with different types of media and media colorant. It follows that it would be useful to supply information concerning media and media colorant to a controller of the printing mechanism so that operation of the printing mechanism could be automatically adjusted to suit the particular media and media colorant. With suitable encryption techniques, this could be used to inhibit after-market refilling. As is well known in the field of printing technology, such after-market refilling has become a cause for substantial concern in the printing industry. In European patent number EP 0779 497, which claims priority from U.S. application Ser. No. 08/573,100, incorporated herein by reference, there is disclosed a fingerprint acquisition sensor. The sensor is described as an apparatus for detecting topological variations on an object such as a finger. The apparatus includes an array of sensing elements disposed on a substrate which each have a parasitic capacitance. An insulating material that covers the sensing elements defines a receiving surface. When a finger is placed on the receiving surface, the capacitance of the sensors changes. A fingerprint comprises ridges and valleys. It will be appreciated that when the finger is positioned on the surface, the ridges create a greater change in capacitance than the valleys. Thus, data representing the fingerprint can be generated using suitable algorithmic electronic circuits. The fingerprint sensor is in the form of a ceramic metal oxide semiconductor (CMOS) device. U.S. Pat. No. 6,049,620 also claims priority from U.S. patent application Ser. No. 08/573,100. This patent discloses a fingerprint-sensing device comprising a planar array of closely spaced capacitive sense elements. The sense elements serve to measure a capacitance between the finger and a single electrode in each sense element. In this patent, each electrode or capacitor plate is charged and then a known current source is used to remove an amount of charge from each plate to measure the capacitance. The measured capacitance varies as a function of a distance between the finger surface and the capacitor plate. Thus, the distance between the finger surface and each capacitor plate can be determined. The distance measurements are used to produce a representation of a pattern of ridges on the finger surface. Applicant has identified a manner in which this form of technology can be applied to achieve a means whereby printing cartridges can be provided with suitable identification data. |
<SOH> SUMMARY OF THE INVENTION <EOH>According to a first aspect of the invention, there is provided a printing cartridge that comprises a housing; and an actuating formation that is positioned on the housing and is capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to at least one of: a serial number of the cartridge, a media and a media colorant, so that the capacitive sensors, when actuated, together generate a signal carrying such data. According to a second aspect of the invention, there is provided a method of determining a media colorant of a printing cartridge, the method comprising the step of actuating a number of capacitive sensors within an array of such sensors with an actuating formation positioned on the printing cartridge, the actuating formation representing data relating to a media colorant in the cartridge, so that the capacitive sensors, when actuated, generate a signal carrying data relating to the media colorant. According to a third aspect of the invention, there is provided a printing cartridge that comprises a housing; a media colorant supply arrangement positioned within the housing and containing a supply of media colorant; and an actuating formation positioned on the housing and being capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to the media colorant so that a signal generated by the capacitive sensors, when actuated, carries said data relating to the media colorant. According to a fourth aspect of the invention, there is provided a method of determining media of a printing cartridge, the method comprising the step of actuating a number of capacitive sensors in an array of such sensors with an actuating formation positioned on the printing cartridge, the actuating formation representing data relating to a media in the cartridge, so that the capacitive sensors, when actuated, generate a signal carrying data relating to the media. According to a fifth aspect of the invention, there is provided a printing cartridge that comprises a housing; a media supply arrangement positioned within the housing and containing a supply of media; and an actuating formation positioned on the housing and being capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to the media so that a signal generated by the capacitive sensors, when actuated, carries said data relating to the media. According to a sixth aspect of the invention, there is provided a method of determining media and media colorant of a printing cartridge, the method comprising the step of actuating a number of capacitive sensors in an array of such sensors with an actuating formation positioned on the printing cartridge, the actuating formation representing data relating to the media and the media colorant in the cartridge, so that the capacitive sensors, when actuated, generate a signal carrying data relating to the media and the media colorant. According to a seventh aspect of the invention, there is provided a printing cartridge that comprises a housing; media and media colorant supply arrangements positioned within the housing and containing a supply of media and a supply of media colorant, respectively; and an actuating formation positioned on the housing and being capable of actuating a number of capacitive sensors in an array of such sensors, the actuating formation being configured to represent data relating to the media and the media colorant so that a signal generated by the capacitive sensors, when actuated, carries said data relating to the media and the media colorant. According to an eighth aspect of the invention, there is provided a printing device which comprises a body, a printing cartridge being engageable with the body, the printing cartridge having a housing, a media colorant supply arrangement positioned within the housing and containing a supply of media colorant, an actuator formation being positioned on the housing and representing data relating to the media colorant; a processor positioned in the body to control operation of a media colorant feed mechanism and a printing mechanism; and an array of capacitive sensors positioned in the body and being configured so that predetermined combinations of capacitive sensors in the sensor array, when actuated, generate signals carrying data related to the media colorant, such predetermined combinations of capacitive sensors in the sensor array being actuable by the actuator formation positioned on the housing of the printing cartridge when the printing cartridge is engaged with the body so that the array of capacitive sensors generates a signal carrying said data relating to the media colorant of the printing cartridge. According to a ninth aspect of the invention, there is provided a printing device which comprises a body, a printing cartridge being engageable with the body, the printing cartridge having a housing, a media supply arrangement positioned within the housing and containing a supply of media, an actuating formation being positioned on the housing and representing data relating to the media; a processor positioned in the body to control operation of a media feed mechanism and a printing mechanism; and an array of capacitive sensors positioned in the body and being configured so that predetermined combinations of capacitive sensors in the sensor array, when actuated, generate signals carrying data related to the media, such predetermined combinations of capacitive sensors in the sensor array being actuable by the actuator formation positioned on the housing of the printing cartridge when the printing cartridge is engaged with the body so that the array of capacitive sensors generates a signal carrying said data relating to the media of the printing cartridge. According to a tenth aspect of the invention, there is provided a printing device which comprises a body, a printing cartridge being engageable with the body, the printing cartridge having a housing, media colorant and media supply arrangements positioned within the housing and containing a supply of media and media colorant, an actuating formation being positioned on the housing and representing data relating to the media colorant and the media; a processor positioned in the body to control operation of media colorant and media feed mechanisms and a printing mechanism; and an array of capacitive sensors positioned in the body and being configured so that predetermined combinations of capacitive sensors in the sensor array, when actuated, generate signals carrying data related to the media and the media colorant, such predetermined combinations of capacitive sensors in the sensor array being actuable by the actuator formation positioned on the housing of the printing cartridge when the printing cartridge is engaged with the body so that the array of capacitive sensors generates a signal carrying said data relating to the media and the media colorant. The invention is now described, by way of example, with reference to the accompanying drawings. The specific nature of the following description should not be construed as limiting in any way the broad nature of this summary. |
Testing device |
A test device (21) sits between two or more nodes (20, 22). The nodes (20, 22) communicate in conversations, according to some predetermined protocol. The test device (21), under user control, may introduce jitter, drop packets, create new packets, reroute packets, and reorder packets in the conversations. Particular conversations are detected and tracked by respective virtual state machines (38, 39, 40) within the test device. |
1. Apparatus having first and second ports passing data therebetween according to a first packet-oriented protocol; the apparatus having first means detecting initiation of a first conversation according to a second packet-oriented protocol overlaid upon the first protocol and, responsive thereto, for creating a respective first virtual state machine, the first virtual state machine disposed to delay a packet of the first conversation; the apparatus having second means detecting initiation of a second conversation according to the second protocol and, responsive thereto, for creating a respective second virtual state machine; said second virtual state machine disposed to emit a causal signal upon a predetermined event in the second conversation; the first virtual state machine disposed to release the delayed packet in response to the causal signal. 2. The apparatus of claim 1 wherein the initiation of the second conversation is subsequent to the initiation of the first conversation. 3. A method for use with apparatus having first and second ports passing data therebetween according to a first packet-oriented protocol, and for use with a conversation-oriented second packet-oriented protocol overlaid upon the first protocol, the method comprising the steps of: detecting initiation of a first conversation according to the second protocol; responsive to the detection, creating a respective first virtual state machine; by the first virtual state machine, delaying a packet of the first conversation; detecting initiation of a second conversation according to the second protocol; responsive thereto the detection, creating a respective second virtual state machine; by the second virtual state machine, emitting a causal signal upon a predetermined event in the second conversation; by the first virtual state machine, releasing the delayed packet in response to the causal signal. 4. The method of claim 3 wherein the detection of the second conversation is subsequent to the detection of the first conversation. 5. Apparatus having first and second ports with data passing into the first port and out of the second port according to a packet-oriented protocol; the apparatus having means responsive to arrival of a first packet at the first port for fragmenting the first packet according to a first pattern, and passing said fragments out of the second port; the apparatus having means responsive to arrival of a second packet at the first port for fragmenting the second packet according to a second pattern different from the first pattern, and passing said fragments out of the second port; the apparatus having means responsive to arrival of a third packet at the first port for fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern, and passing said fragments out of the second port. 6. The apparatus of claim 5 further characterized in that data also pass into the second port and out of the first port according to the packet-oriented protocol; the apparatus having means responsive to arrival of a fourth packet at the second port for fragmenting the fourth packet according to the first pattern, and passing said fragments out of the first port; the apparatus having means responsive to arrival of a fifth packet at the second port for fragmenting the fifth packet according to the second pattern, and passing said fragments out of the first port; the apparatus having means responsive to arrival of a sixth packet at the second port for fragmenting the third packet according to the third pattern, and passing said fragments out of the first port. 7. A method for use with apparatus having first and second ports and for use with a packet-oriented protocol, the method comprising the steps of: receiving a first packet at the first port; fragmenting the first packet according to a first pattern; passing said fragments out of the second port; receiving a second packet at the first port; fragmenting the second packet according to a second pattern different from the first pattern; passing said fragments out of the second port; receiving a third packet at the first port; fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern; and passing said fragments out of the second port. 8. The method of claim 7 wherein the first, second, and third packets are received in the order set forth. 9. The method claim 7 further comprising the steps of: receiving a fourth packet at the second port; fragmenting the fourth packet according to the first pattern; passing said fragments out of the first port; receiving a fifth packet at the second port; fragmenting the fifth packet according to the second pattern; passing said fragments out of the first port; receiving a sixth packet at the second port; fragmenting the sixth packet according to the third pattern; and passing said fragments out of the first port. 10. The method of claim 9 wherein the first, second, third, fourth, fifth, and sixth packets are received in the order set forth. 11. Apparatus having first and second ports with data passing into the first port and out of the second port according to a packet-oriented protocol; the apparatus having means responsive to arrival of a first packet at the first port for fragmenting the first packet according to a first pattern, and passing said fragments out of the second port; the apparatus having means responsive to arrival of a second packet at the first port for fragmenting the second packet according to a second pattern different from the first pattern, and passing said fragments out of the second port. 12. The apparatus of claim 11 further comprising means responsive to arrival of a third packet at the first port for fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern, and passing said fragments out of the second port. 13. The apparatus of claim 11 further characterized in that data also pass into the second port and out of the first port according to the packet-oriented protocol; the apparatus having means responsive to arrival of a fourth packet at the second port for fragmenting the fourth packet according to the first pattern, and passing said fragments out of the first port; the apparatus having means responsive to arrival of a fifth packet at the second port for fragmenting the fifth packet according to the second pattern, and passing said fragments out of the first port. 14. The apparatus of claim 13 further comprising means responsive to arrival of a sixth packet at the second port for fragmenting the third packet according to the third pattern, and passing said fragments out of the first port. 15. A method for use with apparatus having first and second ports and for use with a packet-oriented protocol, the method comprising the steps of: receiving a first packet at the first port; fragmenting the first packet according to a first pattern; passing said fragments out of the second port; receiving a second packet at the first port; fragmenting the second packet according to a second pattern different from the first pattern; and passing said fragments out of the second port. 16. The method of claim 15 further comprising the steps of: receiving a third packet at the first port; fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern; and passing said fragments out of the second port. 17. The method of claim 15 wherein the first and second packets are received in the order set forth. 18. The method of claim 16 wherein the first, second and third packets are received in the order set forth. 19. The method claim 15 further comprising the steps of: receiving a fourth packet at the second port; fragmenting the fourth packet according to the first pattern; passing said fragments out of the first port; receiving a fifth packet at the second port; fragmenting the fifth packet according to the second pattern; and passing said fragments out of the first port. 20. The method of claim 19 further comprising the steps of: receiving a sixth packet at the second port; fragmenting the sixth packet according to the third pattern; and passing said fragments out of the first port. 21. The method of claim 19 wherein the first, second, fourth and fifth packets are received in the order set forth. 22. Apparatus having first and second ports passing data therebetween according to a first packet-oriented protocol; the apparatus having first means detecting initiation of a first conversation according to a conversation-oriented second packet-oriented protocol overlaid upon the first protocol and, responsive thereto, for creating a respective first virtual state machine; the apparatus having second means detecting initiation of a second conversation according to the second protocol and, responsive thereto, for creating a respective second virtual state machine; said second virtual state machine disposed to emit a causal signal upon a predetermined event in the second conversation; the first virtual state machine disposed, in response to the causal signal, to perform a manipulation upon a packet of the first conversation, the manipulation selected from the set consisting of delay, drop, reordering, corruption, and duplication. 23. The apparatus of claim 22 wherein the initiation of the second conversation is subsequent to the initiation of the first conversation. 24. The apparatus of claim 22 wherein the first conversation passes audio data and the second conversation passes video data. 25. The apparatus of claim 22 wherein the first conversation passes video data and the second conversation passes audio data. 26. A method for use with apparatus having first and second ports passing data therebetween according to a first packet-oriented protocol, and for use with a conversation-oriented second packet-oriented protocol overlaid upon the first protocol, the method comprising the steps of: detecting initiation of a first conversation according to the second protocol; responsive to the detection, creating a respective first virtual state machine; detecting initiation of a second conversation according to the second protocol; responsive thereto the detection, creating a respective second virtual state machine; by the second virtual state machine, emitting a causal signal upon a predetermined event in the second conversation; by the first virtual state machine, in response to the causal signal, performing a manipulation upon a packet of the first conversation, the manipulation selected from the set consisting of delay, drop, reordering, rewriting, modification, re-encapsulation, corruption, and duplication. 27. The method of claim 26 wherein the detection of the second conversation is subsequent to the detection of the first conversation. 28. The method of claim 26 wherein the first conversation passes audio data and the second conversation passes video data. 29. The method of claim 26 wherein the first conversation passes video data and the second conversation passes audio data. 30. Apparatus having first and second ports passing data therebetween according to a packet-oriented protocol; the apparatus having first means detecting a first packet of a first predetermined type passing from the second port to the first port; the apparatus having second means passing second packets encoded with a first codec from the first port to the second port; the second means responsive to detection by the first means, and responsive to subsequent detection by the second means of a predetermined event relating to said second packets, for translating said second packets from being encoded with said first codec to being encoded with a second codec differing from the first codec, as the second packets pass from the first port to the second port. 31. The apparatus of claim 30 wherein the first packet of the first predetermined type is an RTCP receiver report. 32. The apparatus of claim 30 wherein the predetermined event relating to said second packets is the passing of a second packet that is an RTP packet with an M-bit set. 33. A method for use with apparatus having first and second ports and for use with a packet-oriented protocol, the method comprising the steps of: detecting a first packet of a first predetermined type passing from the second port to the first port; passing second packets encoded with a first codec from the first port to the second port; detecting a predetermined event relating to said second packets; thereafter translating said second packets from being encoded with said first codec to being encoded with a second codec differing from the first codec, as the second packets pass from the first port to the second port. 34. The method of claim 33 wherein the first packet of the first predetermined type is an RTCP receiver report. 35. The method of claim 33 wherein the predetermined event relating to said second packets is the passing of a second packet that is an RTP packet with an M-bit set. 36. The method of claim 33 further comprising the steps of: again detecting a first packet of the first predetermined type passing from the second port to the first port; passing second packets encoded with the second codec from the first port to the second port; again detecting the predetermined event relating to said second packets; thereafter passing the second packets from the first port to the second port without translating them. 37. Apparatus having a port passing data according to a first packet-oriented protocol; the apparatus having first means detecting initiation of a first conversation according to a second packet-oriented protocol overlaid upon the first protocol and, responsive thereto, for creating a respective first virtual state machine, the first virtual state machine disposed to delay a packet of the first conversation; the apparatus having second means detecting initiation of a second conversation according to the second protocol and, responsive thereto, for creating a respective second virtual state machine; said second virtual state machine disposed to emit a causal signal upon a predetermined event in the second conversation; the first virtual state machine disposed to release the delayed packet in response to the causal signal. 38. The apparatus of claim 37 wherein the initiation of the second conversation is subsequent to the initiation of the first conversation. 39. A method for use with apparatus having a ports passing data according to a first packet-oriented protocol, and for use with a conversation-oriented second packet-oriented protocol overlaid upon the first protocol, the method comprising the steps of: detecting initiation of a first conversation according to the second protocol; responsive to the detection, creating a respective first virtual state machine; by the first virtual state machine, delaying a packet of the first conversation; detecting initiation of a second conversation according to the second protocol; responsive thereto the detection, creating a respective second virtual state machine; by the second virtual state machine, emitting a causal signal upon a predetermined event in the second conversation; by the first virtual state machine, releasing the delayed packet in response to the causal signal. 40. The method of claim 39 wherein the detection of the second conversation is subsequent to the detection of the first conversation. 41. Apparatus having a port with data passing through the port according to a packet-oriented protocol; the apparatus having means responsive to arrival of a first packet at the port for fragmenting the first packet according to a first pattern, and passing said fragments out of the port; the apparatus having means responsive to arrival of a second packet at the port for fragmenting the second packet according to a second pattern different from the first pattern, and passing said fragments out of the port; the apparatus having means responsive to arrival of a third packet at the port for fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern, and passing said fragments out of the port. 42. A method for use with apparatus having a port for use with a packet-oriented protocol, the method comprising the steps of: receiving a first packet at the port; fragmenting the first packet according to a first pattern; passing said fragments out of the port; receiving a second packet at the port; fragmenting the second packet according to a second pattern different from the first pattern; passing said fragments out of the port; receiving a third packet at the port; fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern; and passing said fragments out of the port. 43. The method of claim 42 wherein the first, second, and third packets are received in the order set forth. 44. Apparatus having a port with data passing according to a packet-oriented protocol; the apparatus having means responsive to arrival of a first packet at the port for fragmenting the first packet according to a first pattern, and passing said fragments out of the port; the apparatus having means responsive to arrival of a second packet at the port for fragmenting the second packet according to a second pattern different from the first pattern, and passing said fragments out of the port. 45. The apparatus of claim 44 further comprising means responsive to arrival of a third packet at the port for fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern, and passing said fragments out of the port. 46. A method for use with apparatus having a port and for use with a packet-oriented protocol, the method comprising the steps of: receiving a first packet at the port; fragmenting the first packet according to a first pattern; passing said fragments out of the port; receiving a second packet at the port; fragmenting the second packet according to a second pattern different from the first pattern; and passing said fragments out of the port. 47. The method of claim 46 further comprising the steps of: receiving a third packet at the port; fragmenting the third packet according to a third pattern different from the first pattern and different from the second pattern; and passing said fragments out of the port. 48. The method of claim 46 wherein the first and second packets are received in the order set forth. 49. The method of claim 47 wherein the first, second and third packets are received in the order set forth. 50. Apparatus having a port passing data according to a first packet-oriented protocol; the apparatus having first means detecting initiation of a first conversation according to a conversation-oriented second packet-oriented protocol overlaid upon the first protocol and, responsive thereto, for creating a respective first virtual state machine; the apparatus having second means detecting initiation of a second conversation according to the second protocol and, responsive thereto, for creating a respective second virtual state machine; said second viral state machine disposed to emit a causal signal upon a predetermined event in the second conversation; the first virtual state machine disposed, in response to the causal signal, to perform a manipulation upon a packet of the first conversation, the manipulation selected from the set consisting of delay, drop, reordering, modification, corruption, and duplication. 51. The apparatus of claim 50 wherein the initiation of the second conversation is subsequent to the initiation of the first conversation. 52. The apparatus of claim 50 wherein the first conversation passes audio data and the second conversation passes video data. 53. The apparatus of claim 50 wherein the first conversation passes video data and the second conversation passes audio data. 54. A method for use with apparatus having a port passing data according to a first packet-oriented protocol, and for use with a conversation-oriented second packet-oriented protocol overlaid upon the first protocol, the method comprising the steps of: detecting initiation of a first conversation according to the second protocol; responsive to the detection, creating a respective first virtual state machine; detecting initiation of a second conversation according to the second protocol; responsive thereto the detection, creating a respective second virtual state machine; by the second virtual state machine, emitting a causal signal upon a predetermined event in the second conversation; by the first virtual state machine, in response to the causal signal, performing a manipulation upon a packet of the first conversation, the manipulation selected from the set consisting of delay, drop, reordering, rewriting, modification, re-encapsulation, corruption, and duplication. 55. The method of claim 54 wherein the detection of the second conversation is subsequent to the detection of the first conversation. 56. The method of claim 54 wherein the first conversation passes audio data and the second conversation passes video data. 57. The method of claim 54 wherein the first conversation passes video data and the second conversation passes audio data. 58. Apparatus having a port passing data according to a packet-oriented protocol; the apparatus having first means detecting a first packet of a first predetermined type received at the port and transmitted at the port; the apparatus having second means passing second packets encoded with a first codec received at the port and transmitted at the port; the second means responsive to detection by the first means, and responsive to subsequent detection by the second means of a predetermined event relating to said second packets, for translating said second packets from being encoded with said first codec to being encoded with a second codec differing from the first codec, as the second packets are received at the port and transmitted at the port. 59. The apparatus of claim 58 wherein the first packet of the first predetermined type is an RTCP receiver report. 60. The apparatus of claim 58 wherein the predetermined event relating to said second packets is the passing of a second packet that is an RTP packet with an M-bit set. 61. A method for use with apparatus having a port and for use with a packet-oriented protocol, the method comprising the steps of: detecting a first packet of a first predetermined type received at the port and transmitted at the port; passing second packets encoded with a first codec received at the port and transmitted at the port; detecting a predetermined event relating to said second packets; thereafter translating said second packets from being encoded with said first codec to being encoded with a second codec differing from the first codec, as the second packets are received at the port and transmitted at the port. 62. The method of claim 61 wherein the first packet of the first predetermined type is an RTCP receiver report. 63. The method of claim 61 wherein the predetermined event relating to said second packets is the passing of a second packet that is an RTP packet with an M-bit set. 64. The method of claim 61 further comprising the steps of: again detecting a first packet of the first predetermined type received at the port and transmitted at the port; passing second packets encoded with the second codec received at the port and transmitted at the port; again detecting the predetermined event relating to said second packets; thereafter passing the second packets received at the port and transmitted at the port without translating them. |
<SOH> BACKGROUND <EOH>The application relates generally to test equipment and relates more particularly to test equipment for networks, packet data, and communications links. Historically, and more often before the time of the Internet, many networking and communications protocols were proprietary or at least administered by a controlling entity under circumstances limiting the extent to which third parties could attempt to make compatible equipment and software. In the case of a protocol or standard that was tightly controlled by a particular entity (typically a large corporation), it commonly happened that that entity took it upon itself to certify whether particular third-party equipment was or was not “compatible” with the protocol or standard. In the case of a proprietary standard or protocol it might develop that only the proprietary entity had the ability to make equipment intended to be “compatible,” perhaps due to patents or due to an unpublished “standard” known only to the entity. One natural consequence was that compatibility problems and standards-compliance problems occurred relatively rarely, and another was that barriers to entry often reduced or eliminated competition in particular markets. In more recent times, there has been a trend toward “open” standards, perhaps paralleled to some extent by “open-source” and general-public-license software. Some open standards have come to be widely adopted, in some cases providing levels cross-platform connectivity and interoperability which, as recently as twenty-five years ago, would have been difficult to imagine. For example, in recent years it has come to be possible to exchange data between almost any two operating systems with the help of IP and TCP protocols. Likewise it has come to be possible to exchange email between almost any two email systems, regardless of the designer or the underlying operating system and hardware, with the help of the RFC (request for comments) 822 standard. These trends have led to presence of multiple suppliers in some markets and improved performance and reduced prices in some markets. If an IP packet is lost, nothing about the IP standard will detect the loss. Likewise if two IP packets happen to arrive in a different sequence than they were sent, nothing about the IP standard will detect the change in sequence. To the extent that a system designer wishes to detect and deal with lost or reordered packets, there is no choice but to do it at some protocol level above IP. IP packets do have CRC (cyclic redundancy check) checksums and thus corruption of a packet will nearly always be detected within the IP protocol level. It is convenient to define some terms that characterize things that can go wrong in communications networks. These include the following: Packet loss. This is simply the disappearance of a packet that was transmitted or ought to have been transmitted. Some media have mechanisms to recover lost packets (generally with some additional delay.) For purposes of this note, packets recovered by such media are not considered lost. Packet Delay. Delay is the amount of time that elapses between the time a packet is transmitted and the time it is received. There is always delay —the speed of light in a vacuum places a lower limit on how small the packet delay can be. The actual propagation time of a packet across a telecommunication link varies considerably depending on the media. Some media have complex access mechanisms. For example, CSMA controlled media, such as Ethernets, have fairly intricate access procedures that cause delays often larger than the raw propagation time. In addition to propagation time, packets are delayed as the bits exit and enter computer or switch/router interfaces, as packets spend time in various queues in various switching and routing devices, and as software (or hardware) in those devices examines the packets, including any option fields, and deals with them. Packets are sometimes delayed simply because a computer, router, or switch needs to do something else first, such as doing an ARP handshake to obtain the next hop's MAC address or looking up forwarding information. Jitter. Jitter is a measure of the the variation in the packet delay experienced by a number of packets. Some packets may experience an unimpeded ride through the net while others may encounter various delays. Jitter is often, but not always, accompanied by some degree of packet loss. Jitter is important because it has a significant impact on how long software can expect to wait for data to arrive. That, in turn, has an impact on software buffering requirements. And for media streams, such as voice or video, in which late data is unusable, jitter affects algorithms that are used to create elastic shock-absorbing buffers to provide a smooth play-out of the media stream despite the packet jitter. There are various formulas used to compute jitter. These formulas vary depending on the emphasis that one wants to put on more recent versus older transit variations. One protocol level that is overlaid above IP and that deals with lost and reordered packets is TCP (transmission control protocol). With TCP, a connection is agreed upon between two nodes. The connection has an explicit start and (barring some extreme timeout or loss of connectivity) an explicit end. For the duration of the connection, all packets that are sent are numbered and acknowledged. For the sending node, there is an obligation to preserve a copy of each packet sent until it has been acknowledged. (There is, by definition, some upper limit on how many unacknowledged packets will be permitted to exist at any given moment.) For the receiving node, there is a need to keep track of any gap in the numbers of the received packets, so that when a missing packet finally arrives it can be placed in proper sequence relative to those numbered above and below it. (There is also, by definition, some upper limit as to how many out-of-sequence packets may be stored while the missing packet or packets are awaited.) As a matter of terminology these functions (and others, such as congestion avoidance) are carried out by what is called a “TCP stack.” With the above-mentioned trends come potential problems. Any would-be supplier of a node or network device or system (hereinafter often referred to as a “node”) could offer it to the public as supposedly complying with relevant standards or RFCs, with no choke-point controller (such as a large corporation controlling a standard) to block it. This led to a natural concern as to whether a particular node or device or system was, in fact, compliant with the standard or RFC. As described by Jon Postel in RFC 1025 (September 1987): In the early days of the development of TCP and IP, when there were very few implementations and the specifications were still evolving, the only way to determine if an implementation was “correct” was to test it against other implementations and argue that the results showed your own implementation to have done the right thing. These tests and discussions could, in those early days, as likely change the specification as change the implementation. There were a few times when this testing was focused, bringing together all known implementations and running through a set of tests in hopes of demonstrating the N-squared connectivity and correct implementation of the various tricky cases. These events were called “Bake Offs.” With the growth of the Internet, it became impossible to carry out an “N-squared” demonstration in which each of N devices could be tested for interoperability with the other N−1 devices (and indeed with a device of its own kind), due, among other things, to N becoming very large. In the particular case of TCP connectivity, it became clear that while most TCP stacks performed their desired functions fairly reliably when presented with a connection made up of “ordinary” data, some of them did not perform well in the event of connections made up of inputs that, while standards-compliant, were somewhat out of the ordinary. The traditional approaches to these problems include the following: Design reviews. Designers, engineers, and programmers review a design as it is developed, hoping to assure standards compliance. Code reviews. The actual code written by one or more programmers is discussed with additional programmers, for example with a “walk-through” of the program flow. Protocol test suites. A standardized body of data (e.g. a data file) is fed into a system again and again to test the ability of the system to handle the data. An effort is made to include, within this data file, some fairly wide range of possible inputs. Traffic generators. A device may be created that generates traffic (e.g. packets) according to some particular protocol, thereby testing (among other things) the bandwidth of the node being tested as well as its ability to operate for long periods of time (e.g. to test for certain categories of memory leaks). None of these approaches suffices by itself to test fully the standards compliance of a node, and even these approaches taken together do not lead to complete confidence as to standards compliance. A design review or code review, for example, are performed by humans and thus may completely miss something important that was omitted; it is well known that humans are better at catching something that is visibly incorrect than they are at noticing that something is missing entirely. If a design or body of code is simply lacking a way to test for a boundary condition, for example, this is easy to miss. A distinction can also be drawn between systems and code that are deterministic (that always have the same outputs given certain highly predictable inputs) and systems and code that must deal with a variety of inputs at various times (e.g. asynchronous inputs) and that must deal with potential race conditions among various circuits or data flow paths. An example of a deterministic code body is software to generate, say, daily credit card statements in batches. In the universe of computer programmers and software designers and electrical engineers and systems engineers, almost all are competent to create and to review deterministic systems and code sets. But the fraction of this universe composed of persons who are very good at reviewing the latter types of systems (systems with timing issues, race conditions, asynchronous inputs) turns out to be extremely small. Experience suggests that the need for such people far exceeds the supply. There are not enough of them, for example, to do even a small proportion of the design reviews and code reviews that would be needed to for comprehensive standards-compliance reviews of Internet-related products. While protocol test suites are an important part of testing nodes, they cannot test all or even most of the ways in which a node may be improperly designed. Traffic generators can also be important but again are unlikely to detect subtle design errors. Some prior-art known traffic generators do not, for example, lose packets, reorder packets, corrupt packets, modify packets, create new packets, or delay transfer of packets. Prior-art known traffic generators furthermore do not do these things based upon past traffic. Such traffic generators are not designed to act as an intermediary between two nodes, are not designed to receive packets from multiple sources, do not modify packets based on predetermined or user-controlled criteria, or resend packets which have been intercepted and then modified. One prior-art approach is described by Postel (id.): Some tests are made more interesting by the use of a “flakeway.” A flakeway is a purposely flakey gateway. It should have control parameters that can be adjusted while it is running to specify a percentage of datagrams [packets] to be dropped, a percentage of datagrams to be corrupted and passed on, and a percentage of datagrams to be reordered so that they arrive in a different order than sent. While such flakeways have been devised and actually used for limited testing of TCP stacks, their function has been confined to testing a single connection over a single protocol (e.g. TCP). five-tuple Further, as their function has been limited to simple manipulations only within a single protocol, they do not fully test all of the ways in which a node may fail to handle standards-compliant but infrequent events. With some more recent protocols, there are many options and variants which are within the protocol and yet which are not actually implemented in a first wave of node designs, and which may only come to be implemented in later node designs. This gives rise to a concern that a node from among the first wave may function as expected at first, but may fail to function properly as later-designed nodes commence being put into service. As an example, in fairly recent times it has been proposed to communicate voice information over IP (VoIP). Large portions of the Internet, and many nodes on the Internet, predate VoIP and this raises the natural question whether the existing enterprise network can support VoIP with acceptable voice quality. This raises another natural question namely under what conditions voice quality will be impaired. For example, dynamic routing protocols such as BGP can lead to abrupt changes in the routing of packets during a particular voice conversation, raising the question of how the terminal equipment will handle such changes. Protocols used for VoIP permit the use of any of a number of “codecs” (coders/decoders) which define the manner in which analog signals are converted to digital and later converted back to analog; the protocols further permit shifting dynamically (during a particular voice conversation) from one codec to a different codec. Will such shifts be handled properly? Prior-art test devices do not provide full answers to these questions. Yet another problem of long-standing duration arises from the development of standards which define more than one connection running in parallel (simultaneously). Consider a protocol which defines one connection to pass audio data and another connection to pass video data. Each connection, somewhat analogous to TCP, has mechanisms for detection of and dealing with missing packets. (Depending on the type of data being passed, such as audio data, the protocol may not actually bring about a retransmission of a dropped packet but may instead take some other action such as interpolating the audio signal for the interval represented by the dropped packet.) But it is not enough for each connection, taken by itself, to deal with dropped or delayed or out-of-order packets. There is an additional need, in the example of an audio path and a video path, for the rendered video (perceived by a human) and the rendered audio (again perceived by the same human) to be synchronized. Existing test devices do not fully test this requirement and there is a long-standing need for test devices which would fully test this requirement. Yet another problem of long standing arises from the fact that there are whole categories of design mistakes that are simply not detected if the suite of tests is limited to dropped, delayed, corrupted, duplicated or reordered packets. There is a long-standing need for test devices which would detect more nearly all of the possible design mistakes in modem nodes that are intended to be compliant with present-day standards. |
<SOH> SUMMARY OF THE INVENTION <EOH>A test device sits between two or more nodes. The nodes communicate in conversations, according to some predetermined protocol. The test device, under user control, may introduce jitter, drop packets, create new packets, modify packets, reroute packets, and reorder packets in the conversations. Particular conversations are detected and tracked, for example by respective virtual state machines within the test device. The test device is “stateful” with respect to the protocol; actions taken with respect to a present packet can be based upon past traffic. Importantly, actions may be taken with respect to packets in a first particular conversation based upon past traffic in a second contemporaneous or previous second particular conversation. |
Recorder and cassette containing recording medium |
A recording apparatus capable of recording data either in a current-used format and a new format on both of recording medium accommodated in a current-used tape cassette 50B and a new tape cassette 50A is provided. The recording apparatus comprises a first writing prohibition detection switch 12 to be inserted to an discrimination hole 50Ah3 formed on the cassette 50A and receded in accordance with an open/close state of a writing prohibition discrimination hole provided on the current-used cassette, a second writing prohibition detection switch 10 to be receded in accordance with an open/close state of a writing prohibition discrimination hole 50Ah1 provided on an upper cassette 20, and a 11 to be inserted to an discrimination hole 50Ah2 provided on the upper cassette 50A and receded by a side surface of the current-used cassette. A control circuit 5 discriminates a new tape cassette 50A from the current-used tape cassette 50B in accordance with a state of the switches and performs data recording processing in accordance with the discriminated cassette. |
1. A recording apparatus for discriminating whether a set recording medium is in a first cassette for recording data in a first format or a second cassette for recording data in a second format, detecting whether it is possible to record data on said recording medium accommodated in the discriminated cassette, and recording data in a format in accordance with the discriminated cassette on said recording medium when recordable, comprising: a cassette type detection means (11) for detecting whether said first cassette (50B) or said second cassette (50A, 50C and 50D); a first writing prohibition detection means (12) for detecting whether or not to permit to record data in the first format on said recording medium accommodated in said first cassette or said second cassette; a second writing prohibition detection means (10) for detecting whether or not to record data in the second format on said recording medium accommodated in said first cassette or second cassette; a control means (5); and a data recording means (3, 2); wherein said control means discriminates whether a cassette set in the recording apparatus is said first cassette or said second cassette from a detection signal of said cassette type detection means (11), performing data recording on said recording medium in the first format via said data recording means in the case where a detection signal of said first writing prohibition means (12) permits to write data on said recording medium when said first cassette is set, and performing data recording on said recording medium in the second format via said data recording means in the case where a detection signal of said second writing prohibition means (10) permits to write data on said recording medium when said second cassette is set. 2. A recording apparatus as set forth in claim 1, wherein: said first cassette and said second cassette are formed a first hole (50Bh, 50Ah3, 50Ch3 and 50Dh) for indicating whether said first writing prohibition detection means permits to record data in said first format on said recording medium; said first writing prohibition detection means (12) has a switch having a first pin (12a) able to be inserted to said first hole and outputs an “on” or “off” detection signal in accordance with whether said first pin is inserted to said first hole; said first hole is closed when indicating permission of recording data in said first format on said recording medium, so that said first pin (12a) of said first writing prohibition detection means (12) is prohibited to be inserted to said first hole; and said first hole is open when indicating prohibition of recording data in said first format on said recording medium, so that said first pin (12a) of said first writing prohibition detection means (12) is permitted to be inserted to said first hole. 3. A recording apparatus as set forth in claim 2, wherein: said second cassette is formed a second hole (50Ah2) for indicating that it is a second cassette, and said second writing prohibition detection means (10) is formed a third hole (50Ah1) for indicating whether or not to permit to record data in said second format on said recording medium; said first cassette is not formed said second hole at a part where said second hole is formed on said second cassette, and said first cassette is not formed said third hole at a part where said third hole is formed on said second cassette; said second writing prohibition detection means (10) has a switch having a third pin (10a) able to be inserted to said third hole and outputs an “on” or “off” detection signal in accordance with whether said third pin is inserted to said third hole or not; said third hole is closed when indicating permission of recording data in said second format on said recording medium, so that the third pin (10a) of said second writing prohibition detection means (10) is prohibited to be inserted to said third hole; said third hole is open when indicating prohibition of recording data in said second format on said recording medium, so that the third pin (10a) of said second writing prohibition detection means (10) is permitted to be inserted to said third hole; and said control means identifies that said second cassette is set in the recording apparatus when receiving as an input a signal detecting that a second pin (11a) of said cassette type detection means is inserted to said second hole, and identifies that said first cassette is set in the recording apparatus when receiving as an input a signal detecting that said second pin (11a) of said cassette type detection means is not inserted in said second hole, and when a detection signal of said second writing prohibition detection means (10) permits to record data when said second cassette is set, data recording in the second format is performed on said recording medium via said data recording means. 4. A recording apparatus as set forth in claim 2, wherein: said cassette type detection means (11) and said second writing prohibition detection means (10) are first and second light reception elements (21 and 22) arranged by leaving a predetermined space; a light emission element (20) for emitting a light able to be received by said first and second light reception elements (21 and 22) is further provided; said second cassette is formed a light incident hole (50Ca) for receiving an incident light from said light emission element and first and second emission holes (50Ch1 and 50Ch2) formed at positions corresponding to said two light reception elements (21 and 22), and said second cassette comprises a deflection means (50Cma) for deflecting a light introduced from said light emission element to said light incident hole in the direction of said two light reception elements (21 and 22), and a moving means for deflecting the light introduced from said light emission element to said light incident hole in the direction of one of said two light reception elements by moving the deflection means; said first cassette does not have said emission hole (50Ca) and produced to be opaque; and said control means identifies that a second cassette is set in the recording apparatus when either of said first and second light reception elements receives a light, and performs data recording in the second format on said recording medium via said data recording means when said first light receiving element detects light reception. 5. A recording apparatus as set forth in claim 2, wherein: said cassette type detection means (11) and said second writing prohibition detection means (10) are first data recorded on a first address and a second data recorded on a second address in a semiconductor memory installed in said first cassette and said second cassette; a reading means for reading said first address and second address recorded on said semiconductor memory is further provided; and said control means identifies a cassette set in the recording apparatus is a first cassette when said first data is on, and performs recording processing of data in the second format when said second data is on. 6. A recording medium accommodation cassette, having the same dimensions as those of a first recording medium accommodation cassette for accommodating a recording medium to be recorded data in a first format, characterized by comprising: a first discrimination hole provided at the same position as a writing prohibition setting means provided to said first cassette, having a writing prohibition state; and a discrimination means for discriminating from said first cassette and indicating permission and prohibition of writing to said recording medium accommodation cassette. 7. A recording medium accommodation cassette as set forth in claim 6, characterized in that: said discrimination mechanism comprises a second discrimination hole used for discriminating from said first cassette; and a third discrimination hole for indicating prohibition of writing to said recording medium accommodation cassette by being selectively open or closed. 8. A recording medium accommodation cassette as set forth in claim 6, characterized in that: said discrimination means comprises a light incident hole to which a light emitted from the recording apparatus is irradiated, a reflection member for reflecting said emitted light and selectively sliding between a first position and a second position, a first emission hole for emitting said reflected light when said reflection member is slid to said first position, a second emission hole for emitting said reflected light when said reflection member is slid to said second position, and discrimination from said first cassette and permission or prohibition of writing to said recording medium accommodation cassette are indicated by from which of the first and second emission holes said light is emitted. |
<SOH> TECHNICAL FIELD <EOH>The present invention relates to a recording apparatus for recording data to a cassette, such as a magnetic tape cassette accommodating a magnetic tape as a recording medium, and a recording medium accommodation cassette used therefor. The present invention particularly relates to a recording medium for discriminating two types of cassettes for recording data in different formats on a recording medium and performing data recording processing in accordance with the discriminated cassette, and a recording medium accommodation cassette used therefor. |
<SOH> BRIEF DESCRIPTION OF DRAWINGS <EOH>FIG. 1 is a view of the schematic configuration of a recording/reproducing apparatus according to an embodiment of the present invention. FIG. 2A and FIG. 2B are sectional views as an example of a relationship of a partial section of a tape cassette (a current-used tape cassette) accommodating a magnetic tape wherein data is recorded in a current-used format and a detection switch contacting the current-used tape cassette in a first embodiment of the present invention, wherein FIG. 2A is a view showing a state of permitting data writing in the current-used tape cassette, and FIG. 2B is a view showing a state of prohibiting data writing in the current-used tape cassette. FIG. 3A and FIG. 3B are sectional views as an example of a relationship of a partial section of a tape cassette (a new tape cassette) accommodating a magnetic tape 51 wherein data is recorded in a new (second) format and a detection switch contacting the new tape cassette, wherein FIG. 3A shows a state of permitting data writing in the new tape cassette, and FIG. 3B is a view showing a state of prohibiting data writing in the new tape cassette. FIG. 4 is a flowchart showing operations of a first embodiment of the present invention. FIG. 5 is a view showing the schematic configuration of a recording/reproducing apparatus according to a second embodiment. FIG. 6A and FIG. 6B are sectional views showing an example of an arrangement relationship of a partial section of the new tape cassette, a writing prohibition detection switch, a light emission element and light reception elements, wherein FIG. 6A is a view showing a state of permitting data writing in the new tape cassette, and FIG. 6B is a view showing a state of prohibiting data writing in the new tape cassette. FIG. 7A and FIG. 7B are sectional views showing an example of an arrangement relationship of a partial section of the new tape cassette, a writing prohibition detecting switch, a light emission element and light reception elements, wherein FIG. 7A is a view showing a state of permitting data writing to the current-used tape cassette, and FIG. 7B is a view showing a state of prohibiting data writing to the current-used tape cassette. FIG. 8 is a flowchart showing operations of the second embodiment of the present invention. FIG. 9 is a view showing the schematic configuration of a recording/reproducing apparatus according to a third embodiment of the present invention. FIG. 10 is a sectional view of an example of an arrangement of an outer appearance of a new tape cassette accommodating a magnetic tape wherein data is written in a new format (first format) and the writing prohibition detecting switch in the third embodiment of the present invention. FIG. 11 is a view showing data in a specific address in a maker option region on an IC memory in the third embodiment of the present invention. FIG. 12 is a flowchart showing operations of the third embodiment of the present invention. detailed-description description="Detailed Description" end="lead"? |
Preventing virus infection in a computer system |
A method of preventing an electronic file containing a computer virus from infecting a computer system using the Symbian™ operating system, the method comprising the steps of scanning files using an anti-virus application, and if an infected file is identified, maintaining the file in an open non-sharing state, whereby other applications running on the computer system may not operate on an infected file. |
1. A method of preventing an electronic file containing a computer virus from infecting a computer system, the method comprising the steps of: scanning files using an anti-virus application; and if an infected file is identified, maintaining the file in an open non-sharing state, whereby other applications running on the computer system may not operate on an infected file. 2. A method according to claim 1, wherein the operating system of the computer system is Symbian. 3. A method according to claim 1 wherein, when the operating system of the computer system receives a request from an application to access a previously unaccessed file, the anti-virus application passes a file open command to the operating system which includes as an attribute a non-sharing mode. 4. A method according to claim 3, wherein, if a virus scan of a file reveals that the file is not infected, the anti-virus application returns a file close command to the operating system, whereupon the operating system will set the attributes for the file to closed and sharing, and, if a virus scan reveals that the file is infected, no close command will be returned so that file attributes remain as open non-sharing state. 5. A computer device having processing means arranged in use to scan files using an anti-virus application, and if an infected file is identified, to maintain the file in an open non-sharing state, whereby other applications running on the computer device may not operate on an infected file. 6. A data storage medium having stored thereon a computer program for causing a computer device to scan files using an anti-virus application, and if an infected file is identified, to maintain the file in an open non-sharing state, whereby other applications running on the computer system may not operate on an infected file. |
<SOH> BACKGROUND TO THE INVENTION <EOH>Virus infection of computers and computer systems is a growing problem. Recently there have been many high profile examples where computer viruses have spread rapidly around the world causing many millions of pounds worth of damage in terms of lost data and lost working time. Computer viruses are spread in many different ways. Early viruses were spread by the copying of infected files onto floppy disks, and the transfer of the file from the disk onto a previously uninfected computer. When the user tries to open the infected file, the virus is triggered and the computer infected. More recently, viruses have in addition been spread via the Internet, for example using e-mail. In the future it can be expected that viruses will be spread by the wireless transmission of data, for example by communications between mobile communication devices using a cellular telephone network. Various anti-virus applications are available on the market today. These tend to work by maintaining a database of signatures or fingerprints for known viruses. With a “real time” scanning application, when a user tries to perform an operation on a file, e.g. open, save, or copy, the request is redirected to the anti-virus application. If the application has no existing record of the file, the file is scanned for known virus signatures. If a virus is identified in a file, the anti-virus application reports this to the user, for example by displaying a message in a pop-up window. The anti-virus application may then add the identity of the infected file to a register of infected files. Access to the file is denied. When a subsequent operation on the file is requested, the anti-virus application first checks the register to see if the file is infected. If it is infected, the access is denied. If the file is not infected, access is permitted (the anti-virus application may re-check the file if it detects that the file has changed since the previous check was performed). The approach described in the preceding paragraph for preventing access to infected files is relatively complex and requires a detailed understanding of the workings of the operating system. It also requires some modification to the operating system. Whilst this is allowed (to some extent) by the Microsoft Windows™ operating system, providers of other operating systems may be more reticent to allow interference with their operating systems as this in itself presents a potential security risk. In order to overcome some of the problems, devices may be provided with only an “on-demand” anti-virus scanning application. A user must specifically direct the scanner to scan one file or a group of files. As the application does not have direct access to the operating system, it is not possible to redirect subsequent requests to access an infected file to the anti-virus application. The only option to prevent infection therefore are disinfection of an infected file or, if this is not possible or desirable, deletion of the infected file. |
<SOH> SUMMARY OF THE INVENTION <EOH>According to a first aspect of the present invention there is provided a method of preventing an electronic file containing a computer virus from infecting a computer system, the method comprising the steps of: scanning files using an anti-virus application; and if an infected file is identified, maintaining the file in an open non-sharing state, whereby other applications running on the computer system may not operate on an infected file. Embodiments of the present invention present a simple and elegant solution for “locking” infected electronic files. Typically, no modification is required to the operating system providing that the system allows files to be marked as non-sharing, i.e. only one application at a time can access a file (and an application can only access a file once at any given time). The present invention is applicable in particular to the Symbian™ operating system. Preferably, when the operating system of the computer system receives a request from an application to access a previously unaccessed file, the anti-virus application passes a file open command to the operating system which includes as an attribute a non-sharing mode. The operating system maintains a register of accessed files, and the current file is added to the register with the attributes OPEN and NON-SHARING. While the register records these attributes for the file, if another application attempts to access the file an error will be returned to that application by the operating system. If the scan of the file reveals that the file is not infected, the anti-virus application will return a file close command to the operating system, whereupon the operating system will set the attributes in the register for the file to CLOSED and SHARING. However, if a virus is detected in the file, no close command will be returned. The file will therefore remain open and in a non-sharing state. Again, if another application attempts to open the file, an error will be returned to that application. According to a second aspect of the present invention there is provided a computer device having processing means arranged in use to scan files using an anti-virus application, and if an infected file is identified, to maintain the file in an open non-sharing state, whereby other applications running on the computer device may not operate on an infected file. According to a third aspect of the present invention there is provided a data storage medium having stored thereon a computer program for causing a computer device to scan files using an anti-virus application, and if an infected file is identified, to maintain the file in an open non-sharing state, whereby other applications running on the computer system may not operate on an infected file. |
Adjustable wrench |
An adjustable wrench as in FIG. 1 whereas the wrench 3 become a chain wrench and has a hole through the handle 5 for a chain shackle 4 and the Movable jaw member 2 has a pointed tooth 6 and radiused surfaces 7 on the inclined location. The jaw 2 tongue 8 has a recess 9 for a clock spring 10 its ends being fixed by holes 16 to both the movable jaw tongue 8 and the Handle 3 this spring 10 is preloaded so that the movable Jaw 2 forces towards the gripping surface on handle 3, the coiled portion of the spring being accommodated in the jaw tongue recess 9. The upper corner of movable Jaw 2 incorporates a pistol hammer shaped thumb grip 11. The handle edge 12 being a straight edge ruler with metric and imperial measurements 13 imbedded into the sides of the handle. On the handle ends, there is a radiused hexagonal shaped ring spanner 14 which is for imperial and metric bolts. The gripping surfaces 15 of the handle 3 and Jaw 2 grip surface 7 can be fitted with removable jaw covers called soft Jaws. |
1-27. (canceled) 28. An adjustable wrench comprising, a handle means having a first grip surface and a recess portion formed therein adjacent to the said first grip surface, a movable jaw means having a second grip surface and a tongue portion adapted to be accommodated in the said recess portion, pivot means for pivotally connecting said movable jaw means and said handle means together, the said movable jaw means having a recess formed therein for accommodating a clock spring, a coil portion of the said spring being accommodated in the recess of the said movable jaw means throughout an allowable range of movement of the said movable jaw means so as to bias the said movable jaw means to pivot in a first sense which results in a reduction in distance between said first grip surface and said second grip surface, the handle means having connection means adjacent the first grip surface end of the handle means, the connection means adapted to receive at least one of a chain, cord, and strap and shackle attachment. 29. An adjustable wrench as in claim 28, wherein the connection means is a hole in the handle means, 30. An adjustable wrench as in claim 28, wherein the first grip surface finishes as a point which is half a serration. 31. An adjustable wrench as in claim 28, wherein the first grip surface finishes as a radius which extends to meet an edge of the handle. 32. An adjustable wrench as in claim 28, wherein the connection means is formed to the right size to suit a 6 mm and/or, 8 mm and/or 10 mm chain shackle pin. 33. An adjustable wrench as in claim 28, wherein the width of the wrench is made to fit into the gap of a standard size chain shackle. 34. An adjustable wrench as in claim 28, wherein the handle means is formed with a straight edge having at least one of metric and imperial ruler measurements. 35. An adjustable wrench as in claim 28, wherein the end of the handle means which is remote from the grip surfaces is in the form of a ring spanner which is for at least one of imperial and metric differences, this said ring spanner being offset from a straight edge of the handle means. 36. An adjustable wrench as in claim 28, wherein the recessed portion of the handle means is formed with a step for drilling a hole to hook an outer end of the clock spring, this hole being drilled right through the handle. 37. An adjustable wrench as in claim 28, wherein the said movable jaw means has on an inner inclined surface of the second grip surface a set of curved bumps substantially formed in a cam shape. 38. An adjustable wrench as in claim 28, wherein the said movable jaw means has a radiused portion suitable for receiving a chain shackle pin. 39. An adjustable wrench as in claim 28, wherein an outer part of the movable jaw means incorporates a thumb grip. 40. An adjustable wrench as in claim 28, wherein the clock spring is formed as a coil wound around a pivot pin. 41. An adjustable wrench as in claim 28, wherein the recess of the jaw means is in the tongue portion and is slightly bigger than the thickness of the spring. 42. An adjustable wrench as in claim 28, wherein there is a lip adjacent the outside of the recess in the jaw means which recess is in the tongue portion. 43. An adjustable wrench as in claim 28, wherein there is a hole in the tongue portion for the clock spring to attach to. 44. An adjustable wrench as in claim 28, wherein the clock spring has one end that is bent over approximately 90 degrees to the rest of the spring, this being on an end of the spring that connects to the handle means, and the other end of the spring is also bent over approximately 90 degrees. 45. An adjustable wrench as in claim 28, wherein the said clock spring, adjacent each end part of a coil portion thereof, has a short piece of flat before 90 degree bends at each end. 46. An adjustable wrench as in claim 28, wherein the said clock spring is fitted pretensioned inwards by approximately 45 to approximately 90 degrees. 47. An adjustable wrench as in claim 28, having covers for the first and second grip surfaces. 48. An adjustable wrench as in claim 28, wherein the said handle means comprises at least one of a tapered portion, a lever portion, a folding knife, a spirit level, and a weldable attachment. 49. An adjustable wrench as in claim 28, wherein the said handle means meets the jaw means at an angle of substantially 45 degrees. 50. An adjustable wrench as in claim 28 incorporating at least one of a chain, cord and strap arrangement fitted to the connection means. 51. An adjustable wrench comprising, a handle means having a first grip surface and a recess portion formed therein adjacent to the said first grip surface, a movable jaw means having a second grip surface and a tongue portion adapted to be accommodated in the said recess portion, pivot means for pivotally connecting said movable jaw means and said handle means together, the said movable jaw means having a recess formed therein for accommodating a clock spring, a coil portion of the said spring being accommodated in the recess of the said movable jaw means throughout an allowable range of movement of the said movable jaw means so as to bias the said movable jaw means to pivot in a first sense which results in a reduction in distance between said first grip surface and said second grip surface, the handle means having an aperture adjacent the first grip surface end of the handle means, the aperture adapted to receive at least one of a chain, cord, and strap and shackle attachment. |
<SOH> BACKGROUND <EOH>Wrenches are well known, but many suffer from an undue restriction in terms of the size of objects that they can used with. This is often due to undue restrictions in the jaw openings, and having to change wrench sizes for larger objects. It was noticed while developing the wrench of the present invention that a chain attachment could be added by putting a hole in the corner of the wrench handle end, this hole lets the wrench, with standard link chain and shackles, fit and turn larger objects than was previously possible. Also conventional wrenches suffered from a poor jaw springing system, the spring being unduly exposed to the elements and a large part of the handle was needed to accommodate the springs, for example leaf springs or coil springs. With an embodiment of the wrench of the present invention a clock spring is used such that it is confined to a minimal area in the wrench handle. Also many conventional wrenches have no way of easily retracting or controlling the moving jaw so a suitable thumb grip was designed and added to some embodiments of the invention. Also, with conventional wrenches jaw gripping areas are inclined to damage the work, so a new cam system grip has been designed for the jaw gripping area of an embodiment of the present invention. Also, many conventional wrenches have plain handles, whereas in some embodiments of the present invention full advantage has been taken of the handle by adding a straight edge ruler with metric and imperial measurements embedded either side. Also, it was noticed that common wrenches tend not to utilize the opposite end of the handle to the wrench end. Common wrenches usually have a hole for hanging the wrench, whereas in some embodiments of the invention we have used standard size ring spanners for convenience. Conventional tools such as those mentioned above suffer from various disadvantages and limitations. It is an object of some embodiments of the present invention to go at least some way towards addressing the forgoing problems, or to at least to provide the public with a useful choice. Further aspects and advantages of the present invention will become apparent from the ensuing description which is given by way of example only. The term “comprising” or similar as used herein is not intended to be limiting—for example it should not be taken to infer that there cannot be additional features or the like present. |
<SOH> BRIEF DESCRIPTION OF DRAWINGS <EOH>A preferred embodiment of the invention will now be described with reference to the accompanying drawings in which, FIG. 1 : is an Adjustable Wrench with chain and shackles attached showing the whole wrench with the features and spring area exposed. FIG. 2 : is a picture of the wrench head and handle end clearly showing the shape of the thumb grip and chain hole and jaw gripping system. FIG. 3 : is a picture as in FIG. 1 where as the wrench has the chain and shackle attachment working on a large round object. FIGS. 4 and 5 : are pictures of the wrench end showing a different method of chain attachment. FIG. 6 : is the wrench chain system being used to dog steel into position. FIG. 7 : shows the wrench jaw and a cross section of the said jaw tongue showing the said spring recess and hole for attaching the spring to the head. FIG. 8 : is a clock spring with a side and end view in its free state. FIG. 9 : is the head with the clock spring fitting in its free state. FIG. 10 : shows a wrench end with a section BB through the handle showing the spring area with the spring in position assembled and in its pre-tensioned state. FIGS. 11 - 18 : show the wrench and the jaw cam gripping system when positioned on different sized hexagons and rounds. FIG. 19 : shows the soft jaws on the wrench handle and movable jaw member. FIGS. 20 and 21 : show the movable jaw soft jaws separate from the wrench in 3D with possible different types and showing the cut out for the chain shackle. FIGS. 22 - 23 : show the handle soft jaws shapes in 3D and possible different types. FIG. 24 : shows the wrench with the chain system and the soft jaws on together showing how the soft jaws are accommodated on the wrench when recessed. FIG. 25 : shows the 100 mm wrench design on the imperial side. FIG. 26 : shows the 100 mm wrench movable jaw and handle member separated on the metric side. FIG. 27 : depicts the 150 mm wrench showing the layout of the imperial side, FIG. 28 shows the 150 mm wrench movable jaw and handle member separated and the metric side. FIG. 29 : is the 200 mm wrench showing the shape and layout of the design of the imperial side. FIG. 30 : is the 200 mm wrench showing the shape and layout of the wench movable jaw and handle member separated on the metric side. FIG. 31 : is the 250 mm wrench showing the shape and layout of the design in a assembled state imperial side. FIG. 32 : is the 250 mm wrench showing the shape and layout of the wrench movable jaw and handle separated the metric side. FIGS. 33 and 34 : show the layout and design of the 300 mm model wrench also with a spirit level incorporated in the handle member. FIG. 35 depicts the 300 mm wrench handle member as a edge view showing the ring spanner width is the same as the wrench bodies width. FIG. 36 : shows the 530 mm wrench with a pogy handle and shows an edge view of the 530 mm wrench with a pogy bar for a handle. FIG. 37 : is a wrench handle that is a knife with a sheath on it. FIGS. 38 and 39 : depict the same knife as in FIG. 37 without the sheath, and the knife in the shape of a bowey knife and an edge view of the knife. FIG. 40 : shows a alternative knife handle shape that is in the shape of a dagger which has a sheath. FIG. 41 : shows the wrench as in FIG. 40 with the sheath on. FIGS. 42 and 43 : show a wrench handle that is slotted out having a folding locking pocket knife incorporated. FIG. 44 : depicts a handle for a wrench that is a torch. FIG. 45 : depicts a wrench handle that is a Jimmy lever that would be shaped to pull nails out. FIG. 46 : shows an edge view of the Jimmy lever. FIGS. 47 and 48 : show this type wrench where the handle is of a conventional shape and may have changeable heads. FIGS. 49 - 51 : show the wrench handle with a head in the shape of an axe or chopper. FIG. 52 : shows a wrench handle that has weldable attachments that would be used for fabricating. FIG. 53 : shows the previously mentioned attachments in use. FIG. 54 : shows another type of weld-on attachment. FIGS. 55 - 56 : show a chain system where the head has been replaced with a hook type link to hook onto the chain. FIGS. 57 - 58 : shows a wrench that has an alternatively shaped pogy for a handle that has holes and slots through it, and FIG. 59 - 62 : shows a wrench handle that is retractable and disassembles to breaks, down to small pieces for storage. detailed-description description="Detailed Description" end="lead"? |
Molecular linkers suitable for crystallization and structural analysis of molecules of interest, method of using same, and methods of purifying g protein-coupled receptors |
A method of crystallizing a molecule-of-interest is disclosed. The method comprises (a) contacting molecules of the molecule-of-interest with at least one type of heterologous molecular linker being capable of interlinking at least two molecules of said molecule-of-interest to thereby form a crystallizable molecular complex of defined geometry; and (b) subjecting said crystallizable molecular complex to crystallization-inducing conditions, thereby generating the crystal containing said molecule-of-interest. |
1. A method of generating a crystal containing a molecule-of-interest, the method comprising: (a) contacting molecules of the molecule-of-interest with at least one type of heterologous molecular linker being capable of interlinking at least two molecules of the molecule-of-interest to thereby form a crystallizable molecular complex of defined geometry; and (b) subjecting said crystallizable molecular complex to crystallization-inducing conditions, thereby generating the crystal containing the molecule-of-interest. 2. The method of claim 1, wherein said at least one type of heterologous molecular linker is selected such that said crystallizable molecular complex formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. 3. The method of claim 1, wherein the molecule-of-interest is a polypeptide. 4. The method of claim 3, wherein said polypeptide is a membrane protein. 5. The method of claim 4, wherein said membrane protein is a G protein coupled receptor. 6. The method of claim 5, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 7. The method of claim 6, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 8. The method of claim 1, wherein said at least one type of heterologous molecular linker includes a region for specifically binding the molecule-of-interest. 9. The method of claim 8, wherein the molecule-of-interest is a G protein coupled receptor and whereas said region for specifically binding the molecule-of-interest comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. 10. The method of claim 9, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 11. The method of claim 9, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 12. The method of claim 9, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 13. The method of claim 9, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 14. The method of claim 9, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 15. The method of claim 14, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 16. The method of claim 8, wherein the molecule-of-interest includes a histidine tag and whereas said region for specifically binding the molecule-of-interest comprises a nickel ion or an antibody specific for said histidine tag. 17. The method of claim 8, wherein the molecule-of-interest includes core streptavidin and whereas said region for specifically binding the molecule-of-interest comprises a biotin moiety or a Strep-tag. 18. The method of claim 8, wherein the molecule-of-interest includes a biotin moiety or a Strep-tag and whereas said region for specifically binding the molecule-of-interest comprises core streptavidin. 19. The method of claim 1, wherein the molecule-of-interest is a G protein coupled receptor and whereas said at least one type of molecular linker comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. 20. The method of claim 19, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 21. The method of claim 9, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 22. The method of claim 19, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 23. The method of claim 19, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 24. The method of claim 19, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 25. The method of claim 24, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 26. The method of claim 1 wherein said at least one type of heterologous molecular linker includes at least two non-covalently bound subunits. 27. The method of claim 26, wherein said at least two non-covalently bound subunits comprise a first subunit comprising a homomultimerizing portion and a metal-binding portion, and a second subunit comprising a portion specifically binding the molecule-of-interest, and a portion specifically binding said first subunit. 28. The method of claim 26, wherein said at least two non-covalently bound subunits comprise a first subunit comprising a homomultimerizing portion and a portion specifically binding the molecule-of-interest, and a second subunit comprising a metal-binding portion, and a portion specifically binding said first subunit. 29. The method of claim 1, wherein said at least one type of heterologous molecular linker includes a molecule selected from the group consisting of a polycyclic molecule, a polydentate ligand, a macrobicyclic cryptand, a polypeptide and a metal. 30. The method of claim 1, wherein said at least one type of heterologous molecular linker comprises core streptavidin. 31. The method of claim 1, wherein said at least one type of heterologous molecular linker is selected so as to define the spatial positioning and orientation of said at least two molecules within said crystallizable molecular complex, thereby facilitating crystallization of the molecule-of-interest. 32. The method of claim 1, wherein said at least one type of heterologous molecular linker includes a hydrophilic region, said hydrophilic region being for facilitating crystallization of the molecule-of-interest. 33. The method of claim 1, wherein said at least one type of heterologous molecular linker includes a conformationally rigid region, said conformationally rigid region being for facilitating crystallization of the molecule-of-interest. 34. The method of claim 1, wherein said at least one type of heterologous molecular linker includes a metal-binding moiety capable of specifically binding a metal atom, said metal atom being capable of facilitating crystallographic analysis of the crystal. 35. The method of claim 34, wherein said metal-binding moiety is a metal binding protein. 36. The method of claim 35, wherein said metal binding protein is metallothionein. 37. The method of claim 1, wherein said at least one type of heterologous molecular linker includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of said crystallizable molecular complex and/or of facilitating said interlinking at least two molecules of the molecule-of-interest. 38. The method of claim 37, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. 39. The method of claim 1, wherein the molecule-of-interest includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of said crystallizable molecular complex, and/or of facilitating said interlinking at least two molecules of the molecule-of-interest. 40. The method of claim 39, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. 41. The method of claim 1, wherein the molecule-of-interest includes a metal-binding moiety capable of specifically binding a metal atom, said metal atom being capable of facilitating crystallographic analysis of the crystal. 42. The method of claim 41, wherein said metal-binding moiety is a metal binding protein. 43. The method of claim 42, wherein said metal binding protein is metallothionein. 44. A method of generating a crystal containing a polypeptide of interest, the method comprising: (a) providing a molecule including the polypeptide of interest and a heterologous multimerization domain being capable of directing the homomultimerization of the polypeptide of interest; (b) subjecting said molecule to homomultimerization-inducing conditions, thereby forming a crystallizable molecular complex; and (c) subjecting said crystallizable molecular complex to crystallization-inducing conditions, thereby generating the crystal containing the polypeptide of interest. 45. The method of claim 44, wherein (a) and (b) are effected concomitantly. 46. The method of claim 44, wherein said heterologous multimerization domain is selected such that said crystallizable molecular complex formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. 47. The method of claim 44, wherein said heterologous multimerization domain includes a hydrophilic region, said hydrophilic region being for facilitating crystallization of the polypeptide of interest. 48. The method of claim 44, wherein said heterologous multimerization domain includes a conformationally rigid region, said conformationally rigid region being for facilitating crystallization of the polypeptide of interest. 49. The method of claim 44, wherein said heterologous multimerization domain is selected so as to define the spatial positioning and orientation of polypeptides of the polypeptide of interest within said crystallizable molecular complex, thereby facilitating crystallization of the polypeptide of interest. 50. The method of claim 44, wherein said heterologous multimerization domain comprises core streptavidin. 51. The method of claim 44, wherein the polypeptide of interest is a G protein coupled receptor and whereas said heterologous multimerization domain comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. 52. The method of claim 51, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 53. The method of claim 52, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 54. The method of claim 51, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 55. The method of claim 51, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 56. The method of claim 5 1, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 57. The method of claim 56, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 58. The method of claim 44, wherein the polypeptide of interest includes a histidine tag and whereas said heterologous multimerization domain comprises a nickel ion or an antibody specific for said histidine tag. 59. The method of claim 44, wherein the polypeptide of interest includes core streptavidin and whereas said heterologous multimerization domain comprises a biotin moiety or a Strep-tag. 60. The method of claim 44, wherein the polypeptide of interest includes a biotin moiety or a Strep-tag and whereas said heterologous multimerization domain comprises core streptavidin. 61. The method of claim 44, wherein the polypeptide of interest and said heterologous multimerization domain are interlinked via a molecular linker. 62. The method of claim 61, wherein at least one of said heterologous multimerization domain and said molecular linker include a hydrophilic region, said hydrophilic region being for facilitating crystallization of the polypeptide of interest. 63. The method of claim 61, wherein at least one of said heterologous multimerization domain and said molecular linker include a conformationally rigid region, said conformationally rigid region being for facilitating crystallization of the polypeptide of interest. 64. The method of claim 61, wherein at least one of said heterologous multimerization domain and said molecular linker is selected so as to define the spatial positioning and orientation of polypeptides of the polypeptide of interest within said crystallizable molecular complex, thereby facilitating crystallization of the polypeptide of interest. 65. The method of claim 61, wherein said at least one molecular linker includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of said crystallizable molecular complex, and/or of facilitating said homomultimerization of the polypeptide of interest. 66. The method of claim 65, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. 67. The method of claim 44, wherein the polypeptide of interest includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of said crystallizable molecular complex, and/or of facilitating said homomultimerization of the polypeptide of interest. 68. The method of claim 67, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. 69. The method of claim 44, wherein said molecule includes a metal-binding moiety capable of specifically binding a metal atom, said metal atom being capable of facilitating crystallographic analysis of the crystal. 70. The method of claim 69, wherein said metal-binding moiety is a metal binding protein. 71. The method of claim 70, wherein said metal binding protein is metallothionein. 72. The method of claim 44, wherein the polypeptide of interest is a membrane protein. 73. The method of claim 72, wherein said membrane protein is a G protein coupled receptor. 74. The method of claim 73, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 75. The method of claim 74, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 76. The method of claim 44, wherein the polypeptide of interest includes a metal-binding moiety capable of specifically binding a metal atom, said metal atom being capable of facilitating crystallographic analysis of the crystal. 77. The method of claim 70, wherein said metal binding moiety is metallothionein. 78. A composition-of-matter comprising at least two molecules of a molecule-of-interest interlinked via a heterologous molecular linker, wherein said heterologous molecular linker is selected so as to define the relative spatial positioning and orientation of said at least two molecules within the composition-of-matter, thereby facilitating formation of a crystal therefrom under crystallization-inducing conditions. 79. The composition-of-matter of claim 78, wherein the molecule-of-interest is a polypeptide. 80. The composition-of-matter of claim 79, wherein said polypeptide is a membrane protein. 81. The composition-of-matter of claim 80, wherein said membrane protein is a G protein coupled receptor. 82. The composition-of-matter of claim 81, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 83. The composition-of-matter of claim 82, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 84. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes at least one region capable of specifically binding said molecule-of-interest. 85. The composition-of-matter of claim 84, wherein said molecule-of-interest is a G protein coupled receptor and whereas said at least one region capable of specifically binding said molecule-of-interest is a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. 86. The composition-of-matter of claim 85, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 87. The composition-of-matter of claim 86, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 88. The composition-of-matter of claim 85, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 89. The composition-of-matter of claim 85, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 90. The composition-of-matter of claim 85, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 91. The composition-of-matter of claim 90, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 92. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes a molecule selected from the group consisting of a polycyclic molecule, a polydentate ligand, a macrobicyclic cryptand, a polypeptide and a metal. 93. The composition-of-matter of claim 78, wherein said molecule-of-interest is a G protein coupled receptor and whereas said heterologous molecular linker comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. 94. The composition-of-matter of claim 93, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 95. The composition-of-matter of claim 94, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 96. The composition-of-matter of claim 93, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 97. The composition-of-matter of claim 93, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 98. The composition-of-matter of claim 93, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 99. The composition-of-matter of claim 98, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 100. The composition-of-matter of claim 78, wherein said heterologous molecular linker comprises core streptavidin. 101. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes at least two non-covalently bound subunits. 102. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes a hydrophilic region, said hydrophilic region being for facilitating crystallization of said molecule-of-interest. 103. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes a conformationally rigid region, said conformationally rigid region being for facilitating crystallization of said molecule-of-interest. 104. The composition-of-matter of claim 78, wherein said heterologous molecular linker is selected such that the composition-of-matter is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. 105. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes a metal-binding moiety capable of specifically binding a metal atom, said metal atom being capable of facilitating crystallographic analysis of the crystal. 106. The composition-of-matter of claim 105, wherein said metal-binding moiety is a metal-binding protein. 107. The composition-of-matter of claim 106, wherein said metal binding protein is metallothionein. 108. The composition-of-matter of claim 78, wherein said heterologous molecular linker includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of the crystallizable composition-of-matter, and/or of facilitating said interlinking of said at least two molecules of a molecule-of-interest. 109. The composition-of-matter of claim 78, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. 110. The composition-of-matter of claim 78, wherein said molecule-of-interest includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of the composition-of-matter, and/or of facilitating said interlinking of said at least two molecules of a molecule-of-interest. 111. The composition-of-matter of claim 110, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. 112. The composition-of-matter of claim 78, wherein said molecule-of-interest includes a metal-binding moiety capable of specifically binding a metal atom, said metal atom being capable of facilitating crystallographic analysis of the crystal. 113. The composition-of-matter of claim 112, wherein said metal-binding moiety is a metal binding protein. 114. The composition-of-matter of claim 113, wherein said metal-binding protein is metallothionein. 115. A nucleic acid construct comprising a polynucleotide segment encoding a chimeric polypeptide including: (a) a first polypeptide region being capable of specifically binding a molecule-of-interest; and (b) a second polypeptide region being capable of specifically binding a metal atom. 116. The nucleic acid construct of claim 115, wherein said molecule-of-interest is a G protein coupled receptor and whereas said chimeric polypeptide comprises SEQ ID NO: 5 or SEQ ID NO: 6. 117. The nucleic acid construct of claim 116, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 118. The nucleic acid construct of claim 117, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 119. The nucleic acid construct of claim 115, wherein said molecule-of-interest is a G protein coupled receptor and whereas said first polypeptide region comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. 120. The nucleic acid construct of claim 119, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 121. The nucleic acid construct of claim 120, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 122. The nucleic acid construct of claim 119, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 123. The nucleic acid construct of claim 119, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 124. The nucleic acid construct of claim 119, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 125. The nucleic acid construct of claim 124, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 126. The nucleic acid construct of claim 115, wherein the molecule-of-interest is a polypeptide. 127. The nucleic acid construct of claim 126, wherein said polypeptide is a membrane protein. 128. The nucleic acid construct of claim 127, wherein said membrane protein is a G protein coupled receptor. 129. The nucleic acid construct of claim 128, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 130. The nucleic acid construct of claim 129, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 131. The nucleic acid construct of claim 115, wherein said second polypeptide region is metallothionein. 132. The nucleic acid construct of claim 115, wherein said chimeric polypeptide is selected such that when combined with molecules of said molecule-of-interest under suitable conditions, said chimeric polypeptide and said molecules form a crystallizable molecular complex which is capable of forming a crystal containing said molecule-of-interest when subjected to crystallization-inducing conditions. 133. The nucleic acid construct of claim 115, wherein said chimeric polypeptide is selected such that when combined with molecules of said molecule-of-interest and said metal atom under suitable conditions, said chimeric polypeptide and said molecules form a crystallizable molecular complex which is capable of forming a crystal containing said molecule-of-interest when subjected to crystallization-inducing conditions. 134. The nucleic acid construct of claim 132, wherein said metal atom facilitates crystallographic analysis of said crystal. 135. The nucleic acid construct of claim 132, wherein said chimeric polypeptide includes a hydrophilic region, said hydrophilic region being for facilitating crystallization of said molecule-of-interest. 136. The nucleic acid construct of claim 132, wherein said chimeric polypeptide includes a conformationally rigid region, said conformationally rigid region being for facilitating crystallization of said molecule-of-interest. 137. The nucleic acid construct of claim 132, wherein said chimeric polypeptide is selected so as to define the spatial positioning and orientation of said molecule-of-interest within said crystallizable molecular complex, thereby facilitating crystallization of said molecule-of-interest. 138. The nucleic acid construct of claim 132, wherein said chimeric polypeptide is selected such that said crystallizable molecular complex formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. 139. The nucleic acid construct of claim 132, wherein said chimeric polypeptide further includes a polypeptide region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of said crystallizable molecular complex, and/or of facilitating said binding of a molecule-of-interest. 140. The nucleic acid construct of claim 139, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin. and biotin. 141. A nucleic acid construct comprising a polynucleotide segment encoding a chimeric polypeptide including: (a) a first polypeptide region being capable of specifically binding a molecule-of-interest; (b) a second polypeptide region being capable of homomultimerization into a complex of defined geometry; and (c) a third polypeptide region being capable of specifically binding a metal atom. 142. The nucleic acid construct of claim 141, wherein said molecule-of-interest is a G protein coupled receptor and whereas said first polypeptide region is selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. 143. The nucleic acid construct of claim 142, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 144. The nucleic acid construct of claim 143, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 145. The nucleic acid construct of claim 142, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 146. The nucleic acid construct of claim 9, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 147. The nucleic acid construct of claim 142, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 148. The nucleic acid construct of claim 147, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 149. The nucleic acid construct of claim 141, wherein said second polypeptide region comprises core streptavidin. 150. The nucleic acid construct of claim 141, wherein said molecule-of-interest is a G protein coupled receptor and whereas said chimeric polypeptide comprises SEQ ID NO: 5 or SEQ ID NO: 6. 151. The nucleic acid construct of claim 150, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 152. The nucleic acid construct of claim 151, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 153. The nucleic acid construct of claim 141, wherein said third polypeptide region comprises metallothionein. 154. The nucleic acid construct of claim 141, wherein the molecule-of-interest is a polypeptide. 155. The nucleic acid construct of claim 154, wherein said polypeptide is a membrane protein. 156. The nucleic acid construct of claim 155, wherein said membrane protein is a G protein coupled receptor. 157. The nucleic acid construct of claim 156, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 158. The nucleic acid construct of claim 157, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 159. The nucleic acid construct of claim 141, wherein said chimeric polypeptide is selected such that when combined with molecules of said molecule-of-interest, said chimeric polypeptide and said molecules form a crystallizable molecular complex of defined geometry which is capable of forming a crystal containing said molecule-of-interest when subjected to crystallization-inducing conditions. 160. The nucleic acid construct of claim 159, wherein said chimeric polypeptide includes a hydrophilic region, said hydrophilic region being for facilitating crystallization of said molecule-of-interest. 161. The nucleic acid construct of claim 159, wherein said chimeric polypeptide includes a conformationally rigid region, said conformationally rigid region being for facilitating crystallization of said molecule-of-interest. 162. The nucleic acid construct of claim 159, wherein said chimeric polypeptide is selected so as to define the spatial positioning and orientation of molecules of said molecule-of-interest within said crystallizable molecular complex, thereby facilitating crystallization of said molecule-of-interest. 163. The nucleic acid construct of claim 159, wherein said chimeric polypeptide is selected such that said crystallizable molecular complex of defined geometry formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. 164. The nucleic acid construct of claim 159, wherein said metal atom facilitates crystallographic analysis of said molecule-of-interest contained in said crystal. 165. The nucleic acid construct of claim 159, wherein said chimeric polypeptide further includes a polypeptide region being capable of functioning as a purification tag, said purification tag being capable of facilitating purification of said crystallizable molecular complex, and/or of facilitating said binding of a molecule-of-interest. 166. The nucleic acid construct of claim 165, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, and core streptavidin. 167. A method of purifying a G protein coupled receptor from a sample containing the G protein coupled receptor, the method comprising subjecting the sample to affinity chromatography using an affinity ligand selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, a molecule defined by SEQ ID NO: 3, and a molecule defined by SEQ ID NO: 4, thereby purifying the G protein coupled receptor. 168. The method of claim 167, wherein said at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. 169. The method of claim 168, wherein said at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of said arrestin molecule. 170. The method of claim 167, wherein said mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. 171. The method of claim 167, wherein said mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. 172. The method of claim 167, wherein said G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. 173. The method of claim 172, wherein said class A G protein coupled receptor is m2 muscarinic cholinergic receptor. 174. The method of claim 167, wherein said affinity ligand includes a region being capable of functioning as a purification tag, said purification tag being capable of facilitating attachment of said affinity ligand to an affinity chromatography matrix. 175. The method of claim 174, wherein said region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. |
<SOH> FIELD AND BACKGROUND OF THE INVENTION <EOH>The present invention relates to molecular linkers suitable for crystallization and structural analysis of molecules of interest, to method of using same, and to methods of purifying G protein coupled receptors (GPCRs). More particularly, the present invention relates to methods of crystallizing membrane proteins and to methods of purifying GPCRs via affinity chromatography using arrestin derived polypeptides. Importance of protein structure determination: The recently fully sequenced human genome, has been found to contain up to 38,000 genes (Venter J C. et al., 2001. Science 291:1304) encoding up to an order of magnitude more protein species. It is evident that the information contained therein holds tremendous potential for furthering the development of practical applications in all fields involving the life sciences. However, most proteins remain to be characterized with respect to their structure and function and, although the transcription profiles of the genes encoding these proteins are currently being determined, such data can yield only limited information. In order to fully harness the potential of the information contained in the complete human genome sequence, it will be necessary to systematically determine the three-dimensional (3D) structure of the proteins encoded therein. The capacity to solve the 3D atomic structure of proteins is proving to be crucial for understanding and regulating their biological functions and, as such, is playing an increasingly vital role in the advancement of biomedical science and biotechnology, in particular in the realm of drug design. The pathogenesis of a very large number of human diseases involves membrane proteins such as GPCRs, as startlingly demonstrated by the fact that a 60% majority of approved drugs elicit their therapeutic effects by selectively targeting members of the GPCR family (GlaxoWellcome, 1996. Nature Suppl. 384:1-5). However, pharmacological treatment of diseases involving GPCRs remains far from optimal and there is thus a critical need for novel and improved GPCR-targeting drugs. As highlighted, for example, by the 3D atomic structure-based development of protease inhibitors employed in the first effective treatment of human immunodeficiency virus (HIV) induced acquired immuno-deficiency syndrome (AIDS) (Wlodawer A. and Vondrasek J., 1998. Annu Rev Biophys Biomol Struct. 27:249), the development of novel and improved membrane protein-targeting drugs, such as GPCR-targeting drugs, can dramatically benefit from the availability of the 3D atomic structure of such drug targets. Other increasingly important applications of protein crystals include their use as catalysts on a commercial scale, in bioremediation and green chemistry applications, and in purification-related applications, such as enantioselective chromatography of pharmaceuticals and high-grade chemicals. In the near future, their utility will further expand to include the purification of protein drugs and the development of adjuvant-less vaccines (Margolin A L. and Navia M A., 2001. Angewandte Chemie International Edition 40:2204). General obstacles to protein crystallization: The bottleneck in determination of novel protein structures has shifted from the collection and interpretation of crystallographic data to the production of large amounts of highly pure protein and the generation of diffraction-grade crystals. Techniques for growing such crystals currently rely substantially on empirical processes for which only general rules of thumb are available and which frequently require adaptations tailored to accommodate the peculiarities of individual proteins. Several factors contribute to the difficulty in obtaining highly ordered protein crystals. Although contacts between crystallized protein molecules are of comparable energy to those between small molecules, the significantly fewer number of intermolecular contacts per molecular weight of crystallized protein molecules renders these contacts very fragile (Carugo O. and Argos P., 1997. Protein Science 6:2261). Furthermore, due to their inherent complexity, protein molecules can assume numerous conformations, a phenomenon which tends to prevent formation of highly ordered crystals. Moreover, aggregated proteins are able to form many different types of intermolecular contacts of which only a restricted number will generate highly ordered crystals. Hence, crystallization conditions must be carefully fine-tuned so as to induce the proper molecular conformation and packing orientation of each molecule accreted during the process of crystallization. Such conditions are difficult to obtain since small variations in physico-chemical parameters, such as pH, ionic strength, temperature or contaminants, will strongly influence the process of crystallization in a way that is unique for each protein due to the diversity of the chemical groups and possible configurations thereof involved in the formation of intermolecular contacts (Giege R. et al., Acta Crystallographica Section D-Biological Crystallography 1994. 50:339; Durbin S D. and Feher G., 1996. Annu Rev Phys Chem. 47:171; Weber P C., Overview of protein crystallization methods , in Macromolecular Crystallography , Pt a. 1997. p. 13-22; Chernov A A., Physics Reports-Review Section of Physics Letters 1997. 288:61; Rosenberger F., Theoretical and Technological Aspects of Crystal Growth 1998. p. 241; Wiencek J M., 1999. Annu Rev Biomed Eng. 1:505). Obstacles to Membrane Protein Crystallization Three dimensional protein structure determination at high resolution represents a particularly difficult challenge for membrane proteins and the number of such proteins that have been crystallized is still small and far behind that of soluble proteins, even though membrane proteins represent up to 40% of the proteins encoded by the human genome (Wallin E. and von Heijne G., 1998. Protein Sci. 7:1029). The crystallization of membrane proteins is particularly difficult due to the fact that, unlike soluble proteins which tend to have hydrophilic surfaces and polar cores, membrane proteins have significant hydrophobic surfaces through which they interact with membrane lipids. Such proteins exist in a quasi-solid state in the membrane and are not readily soluble in either aqueous or apolar environments. The most widely employed approach for solubilization of membrane proteins is the use of detergents interacting with the hydrophobic surfaces of the protein to generate mixed detergent/protein micelles. Solubilized membrane proteins can then be crystallized in an ordered two-dimensional (2D) lattice by reconstitution in an artificial lipid bilayer, allowing 2D structural determination via electron microscopy. While such 2D crystals are relatively easy to obtain, the use of electron microscopy for determining molecular structure suffers from the significant drawback of generating structural information with poor resolution in directions orthogonal to the 2D lattice, thus preventing structural determination at sufficiently high resolutions (Stowell M H. et al., 1998. Curr Opin Struct Biol. 8:595). An additional factor contributing to the difficulty of determining the structure of membrane proteins at high resolution is due to the fact that crystal contacts made between detergent micelles tend to be disordered, resulting in poorly diffracting crystals. Although the use of helical crystals and advanced image processing can obviate some of these drawbacks, it is only with X-ray crystallography of 3D crystals that high resolution determination of 3D protein structure can be achieved. This is essential, for example, to generate detailed pictures of molecular target sites when designing drugs specifically interacting with such sites. In the case of membrane proteins, this is highly desirable since such information can significantly contribute to the design and development of novel drugs for the very large number of diseases whose pathogenesis involves membrane proteins, such as receptors. Such diseases include, for example, cancer, viral diseases such as AIDS, neurological disorders, metabolic illnesses such as diabetes, etc. Prior Art Optimization of Crystallization Conditions High Throughput Techniques High throughput techniques are currently being employed to determine the conditions required for growth of protein crystals. One such approach employs automation to perform large numbers of crystallization trials (Morris, D W. et al., 1989. Biotechniques 7:522; Zuk W M. and Ward K B., 1991. Journal of Crystal Growth 110:148; Heinemann U. et al., 2000. Progress in Biophysics & Molecular Biology 73:347). Such high throughput approaches employ the sparse-matrix protein crystallization method, in which a series of crystallization conditions are tested in parallel, the most promising ones being iteratively refined until crystallization is achieved (Jancarik J. and Kim S H., 1991. Journal of Applied Crystallography 24:409; Cudney B., et al., 1994. Acta Crystallographica Section D-Biological Crystallography 50:414; Hennessy D. et al., 2000. Acta Crystallographica Section D—Biological Crystallography 56:817). However, successful crystallization of membrane proteins via such techniques is highly inefficient due to the high tendency of membrane proteins to denature and/or aggregate during crystallization. Furthermore, such methods, being substantially empirical, present the disadvantages of being both time-consuming and of requiring large amounts of pure protein, a requirement which is generally difficult or expensive to fulfill. One strategy which has been suggested in order to circumvent the disadvantages inherent to such high throughput techniques is to assist the crystallization of molecules which are otherwise difficult or impossible to crystallize by either modifying such molecules so as to facilitate their crystallization, or by crystallizing such molecules in complex with other molecules susceptible to provide an ordered matrix facilitating formation of the basic unit of a crystal lattice. Protein-modification techniques: One approach attempting to improve membrane protein crystal growth and ordering has employed complexation of a protein of interest with antibody fragments prior to crystallization (Hunte C., 2001. FEBS Lett. 504:126-32; Lange C. & Hunte C., 2002. Proc Natl Acad Sci USA. 99:2800-5; Ostermeier C. and Michel H., 1997. Curr Opin Struct Biol. 7:697; Ostermeier C. et al., 1997. Proc Natl Acad Sci USA. 94:10547-53). Another modification based approach has used fusion of proteins to be crystallized to large hydrophobic domains derived from heterologous proteins in an attempt to minimize the overall hydrophobicity of proteins of interest (Prive G. et al., 1994. Biol Crystallogr. D50:375). Yet another approach involves alteration and engineering of crystal unit cell contacts, an example being the crystallization of apoferritin by site-directed mutagenesis of residues involved in the binding of a Co 2+ atom introduced during the crystallization process (Takeda S. et al., 1995. Proteins, 23:548). These approaches, however, have the significant drawback that identifying and creating suitable fusion proteins or engineering residues involved in crystal contacts are ad hoc and very labor intensive procedures requiring much fine tuning for applicability to any given protein. Functionalized lipids: Still another approach has employed binding of functionalized lipids to proteins of interest in an attempt to generate crystalline arrays of such proteins. For example, divalent metal ion-chelated lipids or electrostatically charged lipids have been employed to bind proteins via specific surface histidine residues or via complementarily charged residues, respectively. The use of planar layers of such lipids has been employed to generate 2D crystals (Frey W. et al., Proc Nat Acad Sci. USA 1996 93:4937) which can be studied by electron microscopy, but not by X-ray diffraction, thereby yielding limited structural information in terms of dimensionality and in terms of resolution. A more advanced variant of this approach has utilized lipid nanotubes to generate helical crystals (Wilson-Kubalek, E. et al., Proc. Natl. Acad. Sci. U.S.A. 1998, 95:8040). These crystals, however, can only be used to determine 3D protein structure at low resolution using electron microscopy and thus cannot be employed to solve molecular structure at atomic resolution, as is the case with X-ray crystallography. Thus, all prior art approaches have failed to provide an adequate solution for efficiently generating X-ray diffraction grade crystals of molecules such as membrane proteins. There is thus a widely recognized need for and it would be highly advantageous to have, a method of crystallizing molecules, such as membrane proteins, devoid of the above limitations. |
<SOH> SUMMARY OF THE INVENTION <EOH>According to one aspect of the present invention there is provided a method of generating a crystal containing a molecule-of-interest, the method comprising: (a) contacting molecules of the molecule-of-interest with at least one type of heterologous molecular linker being capable of interlinking at least two molecules of the molecule-of-interest to thereby form a crystallizable molecular complex of defined geometry; and (b) subjecting the crystallizable molecular complex to crystallization-inducing conditions, thereby generating the crystal containing the molecule-of-interest. According to further features in preferred embodiments of the invention described below, the at least one type of heterologous molecular linker is selected such that the crystallizable molecular complex formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. According to still further features in preferred embodiments, the molecule-of-interest is a polypeptide. According to still further features in preferred embodiments, the polypeptide is a membrane protein. According to still further features in preferred embodiments, the membrane protein is a G protein coupled receptor. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes a region for specifically binding the molecule-of-interest. According to still further features in preferred embodiments, the molecule-of-interest is a G protein coupled receptor and the region for specifically binding the molecule-of-interest comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the molecule-of-interest includes a histidine tag and the region for specifically binding the molecule-of-interest comprises a nickel ion or an antibody specific for the histidine tag. According to still further features in preferred embodiments, the molecule-of-interest includes core streptavidin and the region for specifically binding the molecule-of-interest comprises a biotin moiety or a Strep-tag. According to still further features in preferred embodiments, the molecule-of-interest includes a biotin moiety or a Strep-tag and the region for specifically binding the molecule-of-interest comprises core streptavidin. According to still further features in preferred embodiments, the molecule-of-interest is a G protein coupled receptor and the at least one type of molecular linker comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes at least two non-covalently bound subunits. According to still further features in preferred embodiments, the at least two non-covalently bound subunits comprise a first subunit comprising a homomultimerizing portion and a metal-binding portion, and a second subunit comprising a portion specifically binding the molecule-of-interest, According to still further features in preferred embodiments, the at least two non-covalently bound subunits comprise a first subunit comprising a homomultimerizing portion and a portion specifically binding the molecule-of-interest, and a second subunit comprising a metal-binding portion, and a portion specifically binding the first subunit. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes a molecule selected from the group consisting of a polycyclic molecule, a polydentate ligand, a macrobicyclic cryptand, a polypeptide and a metal. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker comprises core streptavidin. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker is selected so as to define the spatial positioning and orientation of the at least two molecules within the crystallizable molecular complex, thereby facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes a hydrophilic region, the hydrophilic region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes a conformationally rigid region, the conformationally rigid region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes a metal-binding moiety capable of specifically binding a metal atom, the metal atom being capable of facilitating crystallographic analysis of the crystal. According to still further features in preferred embodiments, the metal-binding moiety is a metal binding protein. According to still further features in preferred embodiments, the metal binding protein is metallothionein. According to still further features in preferred embodiments, the at least one type of heterologous molecular linker includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable molecular complex and/or of facilitating the interlinking at least two molecules of the molecule-of-interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to still further features in preferred embodiments, the molecule-of-interest includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable molecular complex, and/or of facilitating the interlinking at least two molecules of the molecule-of-interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to still further features in preferred embodiments, the molecule-of-interest includes a metal-binding moiety capable of specifically binding a metal atom, the metal atom being capable of facilitating crystallographic analysis of the crystal. According to still further features in preferred embodiments, the metal-binding moiety is a metal binding protein. According to still further features in preferred embodiments, the metal binding protein is metallothionein. According to another aspect of the present invention there is provided a method of generating a crystal containing a polypeptide of interest, the method comprising: (a) providing a molecule including the polypeptide of interest and a heterologous multimerization domain being capable of directing the homomultimerization of the polypeptide of interest; (b) subjecting the molecule to homomultimerization-inducing conditions, thereby forming a crystallizable molecular complex; and (c) subjecting the crystallizable molecular complex to crystallization-inducing conditions, thereby generating the crystal containing the polypeptide of interest. According to further features in preferred embodiments of the invention described below, steps (a) and (b) are effected concomitantly. According to still further features in preferred embodiments, the heterologous multimerization domain is selected such that the crystallizable molecular complex formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. According to still further features in preferred embodiments, the heterologous multimerization domain includes a hydrophilic region, the hydrophilic region being for facilitating crystallization of the polypeptide of interest. According to still further features in preferred embodiments, the heterologous multimerization domain includes a conformationally rigid region, the conformationally rigid region being for facilitating crystallization of the polypeptide of interest. According to still further features in preferred embodiments, the heterologous multimerization domain is selected so as to define the spatial positioning and orientation of polypeptides of the polypeptide of interest within the crystallizable molecular complex, thereby facilitating crystallization of the polypeptide of interest. According to still further features in preferred embodiments, the heterologous multimerization domain comprises core streptavidin. According to still further features in preferred embodiments, the polypeptide of interest is a G protein coupled receptor and the heterologous multimerization domain comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the polypeptide of interest includes a histidine tag and the heterologous multimerization domain comprises a nickel ion or an antibody specific for the histidine tag. According to still further features in preferred embodiments, the polypeptide of interest includes core streptavidin and the heterologous multimerization domain comprises a biotin moiety or a Strep-tag. According to still further features in preferred embodiments, the polypeptide of interest includes a biotin moiety or a Strep-tag and the heterologous multimerization domain comprises core streptavidin. According to still further features in preferred embodiments, the polypeptide of interest and the heterologous multimerization domain are interlinked via a molecular linker. According to still further features in preferred embodiments, at least one of the heterologous multimerization domain and the molecular linker include a hydrophilic region, the hydrophilic region being for facilitating crystallization of the polypeptide of interest. According to still further features in preferred embodiments, at least one of the heterologous multimerization domain and the molecular linker include a conformationally rigid region, the conformationally rigid region being for facilitating crystallization of the polypeptide of interest. According to still further features in preferred embodiments, at least one of the heterologous multimerization domain and the molecular linker is selected so as to define the spatial positioning and orientation of polypeptides of the polypeptide of interest within the crystallizable molecular complex, thereby facilitating crystallization of the polypeptide of interest. According to still further features in preferred embodiments, the at least one molecular linker includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable molecular complex, and/or of facilitating the homomultimerization of the polypeptide of interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to still further features in preferred embodiments, the polypeptide of interest includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable molecular complex, and/or of facilitating the homomultimerization of the polypeptide of interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to still further features in preferred embodiments, the molecule includes a metal-binding moiety capable of specifically binding a metal atom, the metal atom being capable of facilitating crystallographic analysis of the crystal. According to still further features in preferred embodiments, the metal-binding moiety is a metal binding protein. According to still further features in preferred embodiments, the metal binding protein is metallothionein. According to still further features in preferred embodiments, the polypeptide of interest is a membrane protein. According to still further features in preferred embodiments, the membrane protein is a G protein coupled receptor. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the polypeptide of interest includes a metal-binding moiety capable of specifically binding a metal atom, the metal atom being capable of facilitating crystallographic analysis of the crystal. According to still further features in preferred embodiments, the metal binding moiety is metallothionein. According to yet another aspect of the present invention there is provided a composition-of-matter comprising at least two molecules of a molecule-of-interest interlinked via a heterologous molecular linker, wherein the heterologous molecular linker is selected so as to define the relative spatial positioning and orientation of the at least two molecules within the composition-of-matter, thereby facilitating formation of a crystal therefrom under crystallization-inducing conditions. According to further features in preferred embodiments of the invention described below, the molecule-of-interest is a polypeptide. According to still further features in preferred embodiments, the polypeptide is a membrane protein. According to still further features in preferred embodiments, the membrane protein is a G protein coupled receptor. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the heterologous molecular linker includes at least one region capable of specifically binding the molecule-of-interest. According to still further features in preferred embodiments, the molecule-of-interest is a G protein coupled receptor and the at least one region capable of specifically binding the molecule-of-interest is a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the heterologous molecular linker includes a molecule selected from the group consisting of a polycyclic molecule, a polydentate ligand, a macrobicyclic cryptand, a polypeptide and a metal. According to still further features in preferred embodiments, the molecule-of-interest is a G protein coupled receptor and the heterologous molecular linker comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the heterologous molecular linker comprises core streptavidin. According to still further features in preferred embodiments, the heterologous molecular linker includes at least two non-covalently bound subunits. According to still further features in preferred embodiments, the heterologous molecular linker includes a hydrophilic region, the hydrophilic region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the heterologous molecular linker includes a conformationally rigid region, the conformationally rigid region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the heterologous molecular linker is selected such that the composition-of-matter is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. According to still further features in preferred embodiments, the heterologous molecular linker includes a metal-binding moiety capable of specifically binding a metal atom, the metal atom being capable of facilitating crystallographic analysis of the crystal. According to still further features in preferred embodiments, the metal-binding moiety is a metal-binding protein. According to still further features in preferred embodiments, the metal binding protein is metallothionein. According to still further features in preferred embodiments, the heterologous molecular linker includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable composition-of-matter, and/or of facilitating the interlinking of the at least two molecules of a molecule-of-interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to still further features in preferred embodiments, the molecule-of-interest includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the composition-of-matter, and/or of facilitating the interlinking of the at least two molecules of a molecule-of-interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to still further features in preferred embodiments, the molecule-of-interest includes a metal-binding moiety capable of specifically binding a metal atom, the metal atom being capable of facilitating crystallographic analysis of the crystal. According to still further features in preferred embodiments, the metal-binding moiety is a metal binding protein. According to still further features in preferred embodiments, the metal-binding protein is metallothionein. According to still another aspect of the present invention there is provided a nucleic acid construct comprising a polynucleotide segment encoding a chimeric polypeptide including: (a) a first polypeptide region being capable of specifically binding a molecule-of-interest; and (b) a second polypeptide region being capable of specifically binding a metal atom. According to further features in preferred embodiments of the invention described below, the molecule-of-interest is a G protein coupled receptor and the chimeric polypeptide comprises SEQ ID NO: 5 or SEQ ID NO: 6. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the molecule-of-interest is a G protein coupled receptor and the first polypeptide region comprises a molecule selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the molecule-of-interest is a polypeptide. According to still further features in preferred embodiments, the polypeptide is a membrane protein. According to still further features in preferred embodiments, the membrane protein is a G protein coupled receptor. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the second polypeptide region is metallothionein. According to still further features in preferred embodiments, the chimeric polypeptide is selected such that when combined with molecules of the molecule-of-interest under suitable conditions, the chimeric polypeptide and the molecules form a crystallizable molecular complex which is capable of forming a crystal containing the molecule-of-interest when subjected to crystallization-inducing conditions. According to still further features in preferred embodiments, the chimeric polypeptide is selected such that when combined with molecules of the molecule-of-interest and the metal atom under suitable conditions, the chimeric polypeptide and the molecules form a crystallizable molecular complex which is capable of forming a crystal containing the molecule-of-interest when subjected to crystallization-inducing conditions. According to still further features in preferred embodiments, the metal atom facilitates crystallographic analysis of the crystal. According to still further features in preferred embodiments, the chimeric polypeptide includes a hydrophilic region, the hydrophilic region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the chimeric polypeptide includes a conformationally rigid region, the conformationally rigid region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the chimeric polypeptide is selected so as to define the spatial positioning and orientation of the molecule-of-interest within the crystallizable molecular complex, thereby facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the chimeric polypeptide is selected such that the crystallizable molecular complex formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. According to still further features in preferred embodiments, the chimeric polypeptide further includes a polypeptide region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable molecular complex, and/or of facilitating the binding of a molecule-of-interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. According to a further aspect of the present invention there is provided a nucleic acid construct comprising a polynucleotide segment encoding a chimeric polypeptide including: (a) a first polypeptide region being capable of specifically binding a molecule-of-interest; (b) a second polypeptide region being capable of homomultimerization into a complex of defined geometry; and (c) a third polypeptide region being capable of specifically binding a metal atom. According to further features in preferred embodiments of the invention described below, the molecule-of-interest is a G protein coupled receptor and the first polypeptide region is selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, SEQ ID NO: 3, and SEQ ID NO: 4. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the second polypeptide region comprises core streptavidin. According to still further features in preferred embodiments, the molecule-of-interest is a G protein coupled receptor and the chimeric polypeptide comprises SEQ ID NO: 5 or SEQ ID NO: 6. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the third polypeptide region comprises metallothionein. According to still further features in preferred embodiments, the molecule-of-interest is a polypeptide. According to still further features in preferred embodiments, the polypeptide is a membrane protein. According to still further features in preferred embodiments, the membrane protein is a G protein coupled receptor. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the chimeric polypeptide is selected such that when combined with molecules of the molecule-of-interest, the chimeric polypeptide and the molecules form a crystallizable molecular complex of defined geometry which is capable of forming a crystal containing the molecule-of-interest when subjected to crystallization-inducing conditions. According to still further features in preferred embodiments, the chimeric polypeptide includes a hydrophilic region, the hydrophilic region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the chimeric polypeptide includes a conformationally rigid region, the conformationally rigid region being for facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the chimeric polypeptide is selected so as to define the spatial positioning and orientation of molecules of the molecule-of-interest within the crystallizable molecular complex, thereby facilitating crystallization of the molecule-of-interest. According to still further features in preferred embodiments, the chimeric polypeptide is selected such that the crystallizable molecular complex of defined geometry formed is capable of generating a crystal selected from the group consisting of a 2D crystal, a helical crystal and a 3D crystal. According to still further features in preferred embodiments, the metal atom facilitates crystallographic analysis of the molecule-of-interest contained in the crystal. According to still further features in preferred embodiments, the chimeric polypeptide further includes a polypeptide region being capable of functioning as a purification tag, the purification tag being capable of facilitating purification of the crystallizable molecular complex, and/or of facilitating the binding of a molecule-of-interest. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, and core streptavidin. According to a yet a further aspect of the present invention there is provided a method of purifying a G protein coupled receptor from a sample containing the G protein coupled receptor, the method comprising subjecting the sample to affinity chromatography using an affinity ligand selected from the group consisting of at least a portion of an arrestin molecule, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin, at least a portion of an arrestin molecule having a mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin, a molecule defined by SEQ ID NO: 3, and a molecule defined by SEQ ID NO: 4, thereby purifying the G protein coupled receptor. According to further features in preferred embodiments of the invention described below, the at least a portion of an arrestin molecule is homologous to amino acid residues 11 to 190, or 11 to 370 of human beta-arrestin-1a. According to still further features in preferred embodiments, the at least a portion of an arrestin molecule comprises a G protein coupled receptor-binding domain of the arrestin molecule. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 90 in bovine visual arrestin is a mutation to a serine or threonine residue. According to still further features in preferred embodiments, the mutation at an amino acid residue position corresponding to position 175 in bovine visual arrestin is a mutation to a glutamic acid or an asparagine residue. According to still further features in preferred embodiments, the G protein coupled receptor is rhodopsin or is a class A G protein coupled receptor. According to still further features in preferred embodiments, the class A G protein coupled receptor is m2 muscarinic cholinergic receptor. According to still further features in preferred embodiments, the affinity ligand includes a region being capable of functioning as a purification tag, the purification tag being capable of facilitating attachment of the affinity ligand to an affinity chromatography matrix. According to still further features in preferred embodiments, the region being capable of functioning as a purification tag is selected from the group consisting of a T7 tag, a histidine tag, a Strep-tag, core streptavidin, and biotin. |
Assay |
A process for identification of type-specific polynucleotide sequences in a sample, the process comprising the steps of (1) contacting polynucleotides from the sample, or derived from the sample, with a plurality of type-specific probes in the context of a solid support, and detection of any type-specific hybridisation; and (2) contacting polynucleotides in the sample, or derived from the sample, with type-specific primers for those types capable of being detected by the hybridisation step in step 1, but not so detected, in a type-specific amplification reaction. |
1. A process for identification of type-specific polynucleotide sequences in a sample, the process comprising the steps of: (i) contacting polynucleotides from the sample, or derived from the sample, with a plurality of type-specific probes in the context of a solid support, and detection of any type-specific hybridisation; and (ii) contacting polynucleotides in the sample, or derived from the sample, with type-specific primers for those types capable of being detected by the hybridisation step in step 1, but not so detected, in a type-specific amplification reaction. 2. A process according to claim 1 additionally comprising a first step of amplifying polynucleotides from the sample using broad spectrum primers and subsequent use of said amplified polynucleotides in step (i) 3. A process according to claim 2, wherein polynucleotide sequences amplified by broad spectrum primers are analysed to confirm the presence of general polynucleotide types of interest prior to hybridisation. 4. A process according to claim 1 wherein the type specific amplification is quantitative. 5. A process according to claim 1 wherein the type specific primers are specific for one of HPV 16, 18, 31,33, 45, 52, 58, 35, 56, and 59. 6. A method for identification of an individual at risk from disease, the method comprising detecting and typing of a polynucleotide correlated with the disease in a sample from the individual using the process according to claim 1. 7. A diagnostic kit for use in the process of claim 1 comprising at least one set of suitable broad spectrum primers in combination with at least one set of type-specific primers. 8. A kit according to claim 7 comprising HPV 16 and/or HPV 18 type-specific PCR primers. 9. A process, method or kit according to claim 1 for the typing of HPV, HIV-1, HCV, HBV, CMV, EBV, HDV, HGV, HSV, HHV, VZV, Helicobacterpylori, Mycobacteria, mycoplasma, rotavirus or gene p53. 10. A process, method or kit according to claim 9 for the typing of HPV. |
Method and system for enhancing solutions to a system of linear equations |
A method of significantly improving the quality of solutions to a system of linear equations. The solution to a system of linear equations is enhanced by: (1) modeling a target medium into a plurality of elements and imposing at least one localized fluctuation into the target medium; (2) measuring an output resulting from at least one localized fluctuation; and (3) processing the measured output to reconstruct a result, determining a correction filter, and applying the correction filter to the result. |
1. A method of enhancing reconstructed images of a scattering medium, comprising: subdividing a first target medium into a plurality of volume elements; assigning a modulation frequency to at least one of the volume elements' optical coefficients; directing energy into the first target medium from at least one source during a period of time; measuring energy emerging from the first target medium through at least one detector; processing the measured energy emerging from the first target medium to reconstruct at least one image; determining a frequency encoded spatial filter (FESF); and applying the FESF to at least one reconstructed image of the first target medium. 2. The method according to claim 1, wherein the modulation frequency is assigned to an absorption coefficient of a volume element. 3. The method according to claim 1, wherein the modulation frequency is assigned to a scattering coefficient of a volume element. 4. The method according to claim 1, wherein the measured energy emerging from the first target medium is processed by employing a perturbation method. 5. The method according to claim 4, wherein the perturbation method employed uses a forward problem solution to reconstruct the tomographic images. 6. The method according to claim 4, wherein the perturbation method employed uses the inverse problem to reconstruct the tomographic images. 7. The method according to claim 1, wherein determining the FESF, comprises: computing the temporal discrete Fourier transform of the reconstructed tomographic images; and processing the computed temporal discrete Fourier transform to determine the amplitude at a modulation frequency associated with its corresponding volume element. 8. The method according to claim 1, wherein the FESF is applied to at least one reconstructed image by performing a simple matrix multiplication. 9. The method according to claim 1, further comprising: directing energy into a second target medium from at least one source during a period of time; measuring energy emerging from the second target medium through at least one detector; processing the measured energy emerging from the second target medium to reconstruct at least one image; and applying the FESF determined from the first target medium to at least one reconstructed image of the second target medium. 10. A system for enhancing reconstructed images of a scattering medium, comprising: means for subdividing a first target medium into a plurality of volume elements; means for assigning a modulation frequency to at least one of the volume elements' optical coefficients; means for directing energy into the first target medium from at least one source during a period of time; means for measuring energy emerging from the first target medium through at least one detector; means for processing the measured energy emerging from the first target medium to reconstruct at least one image; means for determining an FESF; and means for applying the FESF to at least one reconstructed image of the first target medium. 11. The method according to claim 10, wherein the modulation frequency is assigned to an absorption coefficient of a volume element. 12. The method according to claim 10, wherein the modulation frequency is assigned to a scattering coefficient of a volume element. 13. The method according to claim 10, wherein the measured energy emerging from the first target medium is processed by employing a perturbation method. 14. The method according to claim 13, wherein the perturbation method employed uses a forward problem solution to reconstruct the tomographic images. 15. The method according to claim 13, wherein the perturbation method employed uses the inverse problem to reconstruct the tomographic images. 16. The method according to claim 10, wherein determining the FESF, comprises: means for computing the temporal discrete Fourier transform of the reconstructed tomographic images; and means for processing the computed temporal discrete Fourier transform to determine the amplitude at a modulation frequency associated with its corresponding volume element. 17. The method according to claim 10, wherein the FESF is applied to at least one reconstructed image by performing a simple matrix multiplication. 18. The method according to claim 10, further comprising: means for directing energy into a second target medium from at least one source during a period of time; means for measuring energy emerging from the second target medium through at least one detector; means for processing the measured energy emerging from the second target medium to reconstruct at least one image; and means for applying the FESF determined from the first target medium to at least one reconstructed image of the second target medium. 19. A program stored on a computer readable medium and executable by a processor, comprising: instruction code which, when executed by the processor subdivides a first target medium into a plurality of volume elements; instruction code which, when executed by the processor assigns a modulation frequency to at least one of the volume elements' optical coefficients; instruction code which, when executed by the processor directs energy into the first target medium from at least one source during a period of time; instruction code which, when executed by the processor measures energy emerging from the first target medium through at least one detector; instruction code which, when executed by the processor processes the measured energy emerging from the first target medium to reconstruct at least one image; instruction code which, when executed by the processor determines an FESF; and instruction code which, when executed by the processor applies the FESF to at least one reconstructed image of the first target medium. 20. A program, according to claim 19, further comprising: instruction code which, when executed by the processor directs energy into a second target medium from at least one source during a period of time; instruction code which, when executed by the processor measures energy emerging from the second target medium through at least one detector; instruction code which, when executed by the processor processes the measured energy emerging from the second target medium to reconstruct at least one image; and instruction code which, when executed by the processor applies the FESF determined from the first target medium to at least one reconstructed image of the second target medium. 21. A method of enhancing the solution to a system of linear equations, comprising: modeling a first target medium into a plurality of elements; imposing at least one localized fluctuation to the target medium; measuring an output resulting from at least one localized fluctuation; processing the measured output to reconstruct a result; determining a correction filter; and applying the correction filter to the result. |
<SOH> BACKGROUND <EOH>Imaging of a scattering medium relates generally to a modality for generating an image of the spatial distribution of properties (such as the absorption or scattering coefficients) inside a scattering medium through the introduction of energy into the medium and the detection of the scattered energy emerging from the medium. Systems and methods of this type are in contrast to projection imaging systems, such as x-ray. X-ray systems, for example, measure and image the attenuation or absorption of energy traveling a straight line path between the x-ray energy source and a detector, and not scattered energy. Whether energy is primarily highly scattered or primarily travels a straight line path is a function of the wavelength of the energy and medium it is traveling through. Imaging based on scattering techniques permits the use of new energy wavelengths for imaging features of the human body, earth strata, atmosphere and the like that can not be imaged using projection techniques and wavelengths. For example, x-ray projection techniques may be adept at imaging bone structure and other dense objects, but are relatively ineffective at distinguishing and imaging blood oxygenation levels. This is because the absorption coefficient of blood does not vary significantly with blood oxygenation, at x-ray wavelengths. However, infrared energy can identify the spatial variations in blood volume and blood oxygenation levels because the absorption coefficient at these wavelengths is a function of hemoglobin states. Other structures and functions can be identified by variations or changes in the scattering coefficient of tissue exposed to infrared energy, such as muscle tissue during contraction, and nerves during activation. These structures could not be imaged by projection techniques because projection techniques are not effective in measuring variations in scattering coefficients. These measures, obtainable through imaging based on scattering techniques, such as optical tomography, have considerable potential value in diagnosing a broad range of disease processes. A typical system for imaging based on scattered energy measures, includes at least one energy source for illuminating the medium and at least one detector for detecting emerging energy. The energy source is selected so that it is highly scattering in the medium to be imaged. The source directs the energy into the target scattering medium and the detectors on the surface of the medium measure the scattered energy as it exits. Based on these measurements, a reconstructed image of the internal properties of the medium is generated. The reconstruction is typically carried out using “perturbation methods.” These methods essentially compare the measurements obtained from the target scattering medium to a known reference scattering medium; The reference medium may be a physical or a fictitious medium which is selected so that it has properties that areas close as possible to those of the medium to be imaged. Selecting a reference medium is essentially an initial guess of the properties of the target. In the first step of reconstruction, a “forward model” is used to predict what the detector readings would be for a particular source location based on the known internal properties of the reference medium. The forward model is based on the transport equation or its approximation, the diffusion equation, which describes the propagation of photons through a scattering medium. Next, a perturbation formulation of the transport equation is used to relate (1) the difference between the measured and predicted detector readings from the target and reference, respectively, to (2) a difference between the unknown and known internal properties of the target and reference, respectively. This relationship is solved for the unknown scattering and absorption properties of the target. The final distributions of internal properties are then displayed or printed as an image. Imaging systems and methods based on scattering techniques, such as optical tomography systems, provide a means with which to examine and image the internal properties of scattering media, such as the absorption and diffusion or scattering coefficients. However, the aforementioned imaging systems and methods that recover, contrast features of dense scattering media have thus far produced results having at best modest spatial resolution. Strategies for improving image quality are known (e.g., Newton type), but invariably these are computationally intensive and can be quite sensitive to initial starting conditions. Central to the method of image formation in magnetic resonance imaging (MRI) is that there is a one-to-one correspondence between the frequency of the measured induced current and the spatial orientation of the magnetic field gradient. Because the spatial orientation of the magnetic field gradient is known, this correspondence permits a direct assignment of a measured response to the origin of the signal in space. In effect, the physics of the magnetic resonance phenomenon encodes a frequency signature into the measured data that has a known spatial relationship with the target medium. More generally speaking, methods of this type are known as “frequency encoded spatial filtering.” For the foregoing reasons, there is a need for a computationally efficient nonlinear correction method that is capable of significantly improving the quality of solutions to a system of linear equations such as reconstructed images of a scattering medium. |
<SOH> SUMMARY OF THE INVENTION <EOH>The present invention satisfies this need by providing a method and system for image reconstruction and image correction that is computationally efficient and improves the quality of reconstructed images of a scattering medium. In one embodiment of the system and method of the present invention, reconstructed images of a scattering medium are enhanced by: (1) subdividing a target medium into a plurality of volume elements and assigning a modulation frequency to at least one of the volume elements' optical coefficients; (2) directing energy into the target medium from at least one source during a period of time, and measuring energy emerging from the target medium through at least one detector; and (3) processing the measured energy emerging from the target medium to reconstruct at least one image, determining a frequency encoded spatial filter (FESF), and applying the FESF to at least one reconstructed image. In another embodiment of the system and method of the present invention, a solution to a system of linear equations is enhanced by: (1) modeling a target medium into a plurality of elements and imposing at least one localized fluctuation into the target medium; (2) measuring an output resulting from at least one localized fluctuation; and (3) processing the measured output to reconstruct a result, determining a correction filter, and applying the correction filter to the result. The above advantages and features are of representative embodiments only, and are presented only to assist in understanding the invention. It should be understood that they are not to be considered limitations on the invention as defined by the claims, or limitations on equivalents to the claims. For instance, some of these advantages may seem mutually contradictory, in that they cannot be simultaneously implemented in a single embodiment Similarly, some advantages are primarily applicable to one aspect of the invention. Thus, this summary of features and advantages should not be considered dispositive in determining equivalence. Additional features and advantages of the invention will become apparent in the following description, from the drawings, and from the claims. |
Cellular compositions which facilitate engraftment of hematopoietic stem cells while minimizing the risk of gvhd |
The present invention provides for cellular compositions which facilitate engraftment of hematopoietic stem cells from a syngeneic, allogeneic or xenogeneic donor. The cellular compositions of the invention facilitate engraftment while minimizing the risk of graft versus host disease in the graft recipient. According to a preferred embodiment of the invention, a cell composition is provided, which cell composition comprises hematopoietic stem cells, such as CD34+ cells and/or facilitating cells, in combination with αβ TCR+ T cells. The invention also relates to methods of using the cellular compositions of the invention to induce donor specific tolerance in a recipient, thus allowing the transplantation of donor organs, cells and tissues. Also disclosed are methods of treating leukemia and cancer as well as infectious diseases caused by viruses. |
1. A cellular composition which facilitates donor hematopoietic progenitor cell engraftment into a recipient, comprising donor CD34+ hematopoietic cells in combination with donor αβ TCR+ T cells, wherein the CD34+ cells are at a concentration of about≧0.9×106 cells/kg ideal body weight and the αβ TCR+ T cells are at a concentration of about 0.5-1×105 cells/kg ideal body weight. 2. The cellular composition of claim 1 which further provides at least a 50% reduction in severe GVHD in the recipient. 3. The composition of claim 1, wherein the composition is further comprised of γδ TCR+ T cells. 4. The composition of claim 3, wherein the γδ TCR+ T cells are at a cell concentration of about≧1.5×105 cells/kg ideal body weight. 5. The composition of claim 1 or 3, wherein the composition further comprises NK cells. 6. The composition of claim 1, wherein the composition comprises B cells at a dose below 0.5×105 cells/kg ideal body weight. 7. A pharmaceutical composition for facilitating donor hematopoietic progenitor cell engraftment comprising the composition of claim 1, 2, 3, 4, 5 or 6. 8. A method of partially or completely reconstituting a mammalian lymphohematopoietic system comprising administering to the mammal the pharmaceutical composition of claim 7. 9. The method of claim 8 wherein the mammal is a human. 10. The method of claim 8 wherein the mammal suffers from leukemia, lymphoma or any malignancy of hematolymphoid origin. 11. A method of inducing donor-specific tolerance in a mammal in order to facilitate long term engraftment of donor cells, tissues or organs comprising administering to the mammal the pharmaceutical composition of claim 7. 12. A cellular composition which facilitates donor hematopoietic progenitor cell engraftment into a recipient, comprising donor CD34+ hematopoietic cells in combination with donor facilitating cells (CD8+/TCR−) and donor αβ TCR+ T cells, wherein the CD34+ cells are at a concentration of at least about 0.5×106 cells/kg ideal body weight, the αβ TCR+ T cells are at a concentration of at least about 1×104 cells/kg ideal body weight and the facilitating cells are at a concentration of at least about 0.004×106 cells/kg ideal body weight. 13. The cellular composition of claim 12 which further provides at least a 50% reduction in severe GVHD in the recipient. 14. The composition of claim 12, wherein the composition is further comprised of γδ TCR+ T cells. 15. The composition of claim 14, wherein the γδ TCR+ T cells are at a cell concentration of at least about 1.5×105 cells/kg ideal body weight. 16. The composition of claim 12, wherein the composition further comprises NK cells. 17. The composition of claim 1, wherein the composition comprises B cells at a dose less than about 5×104 cells/kg body weight. 18. A pharmaceutical composition for facilitating donor hematopoietic progenitor cell engraftment comprising the composition of claim 12, 13, 14, 15, 16 or 17. 19. A method of partially or completely reconstituting a mammalian lymphohematopoietic system comprising administering to the mammal the pharmaceutical composition of claim 7. 20. The method of claim 19 wherein the mammal is a human. 21. The method of claim 19 wherein the mammal suffers from a disease selected from the group consisting of leukemia, lymphoma, a malignancy of hematolymphoid origin, hemoglobinopathies such as sickle cell anemia, spherocytosis, thalassemia and other blood disorders, autoimmune disease, diabetes, and metabolic disorders such as Hunters disease, Hurlers disease, and enzyme defects. 22. A method of inducing donor-specific tolerance in a mammal in order to facilitate long term engraftment of donor cells, tissues or organs comprising administering to the mammal the pharmaceutical composition of claim 18. 23. The cellular composition of claim 12 wherein the hematopoietic facilitating cells have a phenotype of CD3+, CD8+, αβ TCR− and γδ TCR− as determined by antibody staining and flow cytometry, which hematopoietic facilitating cells are capable of facilitating engraftment of bone marrow cells. 24. The cellular composition of claim 23 wherein the hematopoietic facilitating cells are CD45+. 25. The cellular composition of claim 24 wherein the hematopoietic facilitating cells are CD45R+. 26. The cellular composition of claim 25 wherein the hematopoietic facilitating cells are Thy1+, CD19− and CD56−. 27. The cellular composition of claim 26 wherein the composition comprises at least about 30% human hematopoietic facilitating cells having a phenotype of CD3+, CD8+, αβ TCR− and γδ TCR− as determined by antibody staining and flow cytometry. 28. The cellular composition of claim 26 wherein the composition comprises at least about 95% human hematopoietic facilitating cells having a phenotype of CD3+, CD8+, αβ TCR− and γδ TCR− as determined by antibody staining and flow cytometry. 29. The cellular composition of claim 12 wherein the hematopoietic facilitating cells are capable of facilitating engraftment of bone marrow cells. 30. The cellular composition of claim 29 wherein the hematopoietic facilitating cells are CD45+. 31. The cellular composition of claim 30 wherein the hematopoietic facilitating cells are CD45R+. 32. The cellular composition of claim 31 wherein the hematopoietic facilitating cells are Thy1+, CD19− and CD56−. 33. The cellular composition of claim 29 further comprises CD34+ cells that are histocompatible with the hematopoietic facilitating cells. 34. The cellular composition of claim 12 wherein the hematopoietic facilitating cells are at a concentration of at least about 0.5×106 cells/kg ideal body weight. 35. The cellular composition of claim 34 wherein the CD34+ cells are at a concentration of at least about 0.9×106 cells/kg ideal body weight, the αβ TCR+ T cells are at a concentration of at least about 1×105 cells/kg ideal body weight. 36. The cellular composition of claim 35 wherein the CD34+ cells are at a concentration of at least about 2×106 cells/kg ideal body weight. |
<SOH> 2. BACKGROUND OF THE INVENTION <EOH>The goal of hematopoietic progenitor cell or stem cell transplantation is to achieve the successful engraftment of donor cells within a recipient host such that immune and/or hematopoietic chimerism results. Chimerism is the reconstitution of the various compartments of the recipient's hematoimmune system with donor cell populations bearing major histocompatability complex (MHC) molecules derived from both the allogeneic or xenogeneic donor and a cell population derived from the recipient or alternatively the recipient's hematoimmune system compartments which can be reconstituted with a cell population bearing MHC molecules derived from only the allogeneic or xenogeneic marrow donor. Chimerism may vary from 100% (total replacement by allogenic or xenogeneic cells) to low levels detectable only by molecular methods. Chimerism levels may vary over time and be permanent or temporary. Hematopoietic progenitor cells or stem cells are pluripotent cells that are capable of reconstituting a recipient's hematoinmmune system. Early hematopoietic progenitor cells are characterized by the presence of several cell surface markers, including CD34 (see, e.g., European patent application EP 0 451 611 A2). Reconstitution of the recipient's hematoimmune system is accomplished by transferring a heterogeneous population of cells, including hematopoietic stem or progenitor cells, derived from the donor's bone marrow or peripheral blood to the graft recipient. The challenge to achieving successful donor cell chimerism involves balancing the outcomes of graft rejection, graft versus leukemia (when the graft is performed in the context of treating a patient suffering from leukemia), immune reconstitution and graft versus host disease (GVHD). Graft rejection occurs when the donor cells fail to reconstitute the target compartments of the recipient's hematoimmune system. GVHD occurs when the graft of the donor cells is successful, but the immunocompetent donor cells recognize and attack the recipient's organs and tissues. Graft versus leukemia (GVL) is the recognition and destruction of residual leukemia cells by transplanted immune cells. When the aforementioned circumstances are optimally achieved, the immunological circumstance is generally associated with tolerance. Bone marrow and/or stem cell transplantation has applications in a wide variety of clinical settings, including solid organ transplantation. A major goal in solid organ transplantation is the engraftment of the donor organ without a graft rejection immune response generated by the recipient, while preserving the immunocompetence of the recipient against other foreign antigens. Typically, nonspecific immunosuppressive agents such as cyclosporine, methotrexate, steroids and FK506 are used to prevent host rejection responses. They must be administered on a daily basis and if stopped, graft rejection usually results. However, nonspecific immunosuppressive agents function by suppressing all aspects of the immune response, thereby greatly increasing a recipient's susceptibility to infections and diseases, including cancer. Furthermore, despite the use of immunosuppressive agents, graft rejection still remains a major source of morbidity and mortality in human organ transplantation. It would therefore be a major advance if tolerance can be induced in the recipient. The only known clinical circumstance in which complete systemic donor-specific transplantation tolerance has been induced is allogeneic hematopoietic stem cell transplantation (HSCT). (See Qin et al., 1989, J. Exp. Med. 169:779; Sykes et al., 1988, Immunol. Today 9:23; Sharabi et al., 1989, J. Exp. Med. 169:493). This has been achieved in neonatal and adult animal models as well as in humans by total lymphoid irradiation, total body irradiation or immunosuppressive chemotherapy of a recipient followed by bone marrow transplantation with donor cells. The success rate of HSCT is, in part, dependent on the ability to closely match the MHC of the donor cells with that of the recipient cells. The MHC is a gene complex that encodes a large array of glycoproteins expressed on the surface of both donor and host cells that are vital to normal function of the immune system, but are also the major targets of the transplantation rejection immune response. In humans, MHC is referred to as Human Leukocyte Antigen (HLA). HLA genes are inherited in a Mendelian fashion, hence, the only hope for a donor with an identical set of HLA proteins is in a sibling with the identical inheritance pattern. Transplants from a matched sibling donor are still associated with significant levels of GVHD, but meet with a high degree of success. However, when allogeneic bone marrow transplantation is performed between two MHC-mismatched individuals of the same species, common complications involve failure of engraftment, poor immunocompetence and a high incidence of GVHD. GVHD is a potentially lethal complication in bone marrow transplantation, which occurs in about 35-50% of recipients of untreated HLA-identical marrow grafts (Martin et al., 1985, Blood 66:664) and up to 80% of recipients of HLA-mismatched marrow. Unfortunately, only 30% of patients generally have a suitably matched HLA-identical family member donor, and thus most patients are either excluded from being considered for bone marrow transplantation, or if they are transplanted, must tolerate a high risk of GVHD. GVHD results from the ability of immunocompetent mature immune cells, largely αβ TCR + T cells, in the donor graft to recognize host tissue antigens as foreign and produce an adverse immunologic reaction. Recent studies in bone marrow transplantation suggest that the major cause of GVHD is T cells, as the removal of T cells from the donor cell preparation was associated with a reduction in the incidence of GVHD. (Vallera et al., 1989, Transplant, 47:751; Rayfield, 1984, Eur. J. Immunol., 4:308; Vallera, 1982, J. Immunol., 128:871; Martin and Korngold, 1978, J. Exp. Med., 148(6):1687; Prentice, 1984, Lancet 1(8375): 472). After T cells were implicated as the predominant mediator of GVHD in animal models, aggressive protocols for T cell depletion (TCD) of human donor bone marrow were instituted. Technology now exists to deplete bone marrow of T cells. These techniques include the use of monoclonal antibodies (in conjunction with magnetic beads, immunotoxins or complement lysis), gradient fractionation, and soybean lectin agglutination. (See Kernan, 1994, Bone Marrow Transplantation, Oxford: Blackwell Scientific Publications, p. 124-35). Although TCD decreased the incidence of GVHD dramatically, TCD was accompanied by a significant increase in the failure of engraftment, indicating that T cells might also play a facilitating role in bone marrow engraftment. (Soderling, J. Immunol., 1985, 135:941; Vallera, 1982, Transplant. 33:243; Pierce, 1989, Transplant., 48(2):289). TCD in the context of treating leukemia patients is also associated with an increased risk of leukemia relapse. It is therefore believed that T cells contained in the donor graft are instrumental in mediating this anti-leukemic effect known as GVL (Champlin et al. 1996, Acta Haematol 95: 157). In addition, the infused T cells probably produced immunologic effects against viruses and other pathogens, as evidenced by the increase in opportunistic infections in patients after TCD transplants. The increase in failure of engraftment in human recipients ranges from about 5-70% of total patients and is related to the degree of MHC disparity between the donor and recipient and the breadth and completeness of TCD (Blazer, 1987, UCLA Symp., p. 382; Filipovich, 1987, Transplant., 44(1):62; Martin et al., 1985, Blood 66:664; Martin et al., 1988, Adv. Immunol. 40:379). Patients with failed engraftment usually die even if a second bone marrow transplant is performed. Consequently, most transplant institutions in the United States have abandoned TCD of donor bone marrow and, thus, must tolerate a high level of GVHD, which leads to significant morbidity and mortality. Thus, the application of bone marrow transplantation as a form of treatment is limited only to settings where the potential of GVHD is clearly outweighed by the potential benefit. The implication that T cells might participate in both harmful GVHD reactions and helpful outcomes such as engraftment facilitation, GVL and hematoimmune reconstitution was an enigma that existed for a long time in the scientific community. Investigators began to search for the possible existence of a bone marrow component that could facilitate bone marrow engraftment but was removed during TCD. Identification and purification of this facilitating component would potentially allow the design of transplant protocols to selectively prevent GVHD, while preserving the cells that can enhance engraftment and thus allow the application of allogeneic and xenogeneic bone marrow transplantation to be used in a wide variety of clinical settings. Cell populations that mediate graft facilitating or anti-leukemic effects have been identified in murine models (See U.S. Pat. No. 5,772,994; Kaufman et al., Thirty Fifth Annual Meeting of the American Society of Hematology , St. Louis, Mo., 1993, 82(10 Suppl. 1):456A). 76 TCR + T cells have been shown to possess anti-leukemic activity against ALL (Lamb et al., 2001, Bone Marrow Transplantation 27:601). NK cells mismatched at receptor loci have also been shown to possess anti-leukemic activity (Valiante et al., 1997, Biol. Blood Marrow Transplant 3(5):229). αβ TCR + T cells have been shown to possess suppressive activity against a wide variety of leukemias and other malignancies. Megadoses (>10 7 kg/Ideal Body Weight) of CD34 + hematopoietic stem cells have been shown to have a beneficial effect in achieving engraftment across MHC barriers with little GVHD in some leukemia patients (Reisner et al.,1999, Ann. N.Y. Acad. Sci. 872:336). A subset of CD34 + cells called “veto cells” may induce anti-host unresponsiveness in the infused T cells (Gur et al., 1999, Blood 94(Suppl.):391; Bachar-Lustig et al., 1999, Blood 94:3212). Conversely, doses of 10 5 cells/kg recipient body weight using cord blood have also produced engraftment (Laughlin et al., 2001, N. Eng. J Med 3(5):229). The chimeric immune system must also function to recognize and destroy pathogens. It has also been reported that a distinct cell type bearing cell surface markers including CD8 + , CD3 + , Thy1 + , ClassII dim/intermediate , CD45 + , αβ TCR − T cells, and γδ TCR − T cells can facilitate allogeneic and xenogeneic engraftment with little GVHD (U.S. Pat. No. 5,772,994; Kaufman et al., supra). In addition, studies have suggested that T cells, generally, may play a role in facilitating engraftment (Lapidot et al., 1992, Blood, 80(9):2406; Kernan et al.,1986, Blood, 68(3):770). These studies have not defined specific T cell subsets or doses that may be beneficial in facilitating engraftment. Studies have been done, using a mouse model, which suggest that αβ TCR + T cells and γδ TCR + T cells may facilitate engraftment (Drobyski et al., 1997, Blood, 89(3):1100). However, the predicted beneficial dose ranges of the T cell subsets in this study are not within the ranges provided for by the instant invention. The invention disclosed herein describes compositions of cells isolated from bone marrow and/or peripheral blood and the uses of these cellular compositions to facilitate desired clinical outcomes including bone marrow and/or hematopoietic progenitor cell engraftment by minimizing the risk of both graft rejection and GVHD, while allowing for hematopoietic and/or immune reconstitution. In the context of treating leukemia, the cellular compositions of the invention maintain the graft versus leukemia effect. |
<SOH> 3. SUMMARY OF THE INVENTION <EOH>The present invention provides cellular compositions and methods of treating a host mammal in need of a hematopoietic progenitor cell transplant by identifying cellular populations that facilitate achieving desirable outcomes such as engraftment of hematopoietic stem cells. According to a preferred embodiment of the invention, a cell composition is provided, which cell composition comprises hematopoietic progenitor cells, such as CD34 + cells, in combination with αβ TCR + T cells. The cells utilized in the cellular compositions of the invention may be derived from any physiological source, for example, but not as a limitation, bone marrow. As noted in Section 2, supra, hematopoietic progenitor cells carrying the CD34 marker, when administered in large numbers, appear to correlate with a reduced time to engraftment (Reisner et al., supra). Conversely, the use of low numbers of CD34 + cells correlates with much longer times to engraftment. However, obtaining large numbers of CD34 + early progenitor cells can be difficult. The present invention allows for the use of smaller quantities of CD34 + cells in allogeneic hematopoietic stem cell transplants by combining the hematopoietic progenitor cells with certain populations of cells that facilitate engraftment. Concentrations of CD34 + cells according to the compositions and methods of this invention can be in the range of about 0.5×10 6 to about 2.5×10 6 cells/kg ideal body weight (IBW), preferably in the range of about 0.9×10 6 to about 2.0×10 6 cells/kg ideal body weight (COW). More specifically, applicants have discovered that T cells that carry the of TCR marker (referred to herein as αβ TCR + T cells) are correlated with producing a shorter time to engraftment or both neutrophils and platelets when they are present in bone marrow transplants. As exemplified infra in Section 6.12, this subset of T cells correlates with producing shorter times to engraftment using a univariate and/or multivariate mathematical analysis. Thus, in one embodiment, the invention comprises a cellular composition comprising CD34 + hematopoietic cells and αβ TCR + T cells. T cells that carry the γδ TCR marker (referred to herein as γδ TCR + T cells) also show a tendency to produce shorter times to engraftment of both platelets and neutrophils and they are not correlated with producing acute GVHD. Accordingly, an alternative embodiment of the present invention comprises a cellular composition comprising CD34 + hematopoietic cells, αβ TCR + T cells, and γδ TCR + T cells. Another embodiment of the present invention comprises a cellular composition comprising CD34 + hematopoietic cells, facilitating cells, αβ TCR + T cells, and γδ TCR + T cells. In addition, since NK cells correlate with producing shorter times to engraftment in a univariate statistical model (See Section 6.12 infra), still another embodiment of the invention comprises any of the cellular compositions described above in combination with NK cells. Finally, B cells do not appear to facilitate engraftment and are a source of post-transplant complication, particularly post-transplant lymphoproliferative disorder. Therefore, according to one embodiment of the invention, the number of B cells in hematopoietic stem cell transplantation is limited. The invention therefore relates to cellular compositions comprising CD34 + cells in combination with one or more of the following cell populations: αβ TCR + T cells, γδ TCR + T cells, NK cells and/or B cells in appropriate concentrations such that host engraftment is facilitated while the risk of inducing GVHD and/or other post-transplant complications is minimized. Preferably, the cellular compositions comprising CD34 + cells in combination with facilitating cells along with one or more of the following cell populations: αβ TCR + T cells, γδ TCR + T cells, NK cells and/or B cells in appropriate concentrations such that host engraftment is facilitated while the risk of inducing GVHD and/or other post-transplant complications is minimized. In promoting hematopoietic engraftment in a host that may be HLA-mismatched, including, for example, allogeneic, or xenogeneic transplants, the invention may be used for the generation of donor tolerance by the host. Coexistence of donor and host marrow or complete replacement of host marrow by donor cells and the subsequent creation of a mammalian chimera by the invention provides a new method for generating tolerance to either solid organ or tissue transplants. Transplantation of donor hematopoietic progenitor cells allows the subsequent and/or simultaneous transplantation of solid organs or tissues from the same donor such as but not limited to heart, kidney and liver. The recipients' new immune system recognizes the transplanted organs as ‘self’ as opposed to ‘non-self.’ Thus, the invention provides methods for treating a host mammal with an allogeneic or xenogeneic hematopoietic progenitor cell preparation to allow engraftment of the new cell material into the host. The host mammal usually undergoes immunosuppression before administration of the transplant of hematopoietic cells. Immunosuppression of the host reduces its ability to recognize and reject the donor cells of the graft. Pharmacologic methods, immunologic methods and/or irradiation may achieve this immunosuppression of the host, referred to herein as conditioning, for example. By utilizing the cell compositions and methods of the invention, the clinical outcome of the recipient prepared by various conditioning approaches can be enhanced. The clinical outcomes or endpoints that can be influenced include but are not limited to: (i) serious acute graft versus host disease (defined as grades 3 and 4), (ii) chronic graft versus host disease, (iii) post-transplant lymphoproliferative disease, (iv) engraftment of both platelets and neutrophils, (v) immune reconstitution, (vi) disease relapse (examples include leukemia, lymphoma and sickle cell disease), (vii) overall survival or (viii) tolerance. The compositions and methods of this invention can be used in the treatment of any disease or condition requiring a hematopoietic progenitor or stem cell transplant. By way of example, but not as a limitation, such diseases can include: cancer, leukemia, lymphoma, or any malignancy of hematolymphoid origin, autoimmune disease, AIDS, any disease resulting in immunodeficiency, sickle cell disease, anemia, diabetes or any viral infection. The compositions may also be used to treat a recipient in need of an organ or tissue transplant. Non-limiting examples of organ transplants for which the compositions might be used include heart, lung, liver, kidney, pancreas or skin graft. The compositions may also be used in tissue transplants such as islet cells of the pancreas or dopamine-producing brain cells. The examples provided are not intended to be limiting and one skilled in the art would appreciate that the invention would have application in any clinical setting where transplantation is used. |
Semiconductor device and method of its manufacture |
The present invention presents a semiconductor device (10) which is adapted to a solar cell, and in which a semiconductor element (1) is produced by forming one flat surface (2) on a spherical or substantially spherical silicon single crystal (1a, 1b). A diffusion layer (3) and a substantially spherical pn junction (4) are formed on this semiconductor element (1), and a diffusion-mask thin film (5) and a positive electrode (6a) are formed on the flat surface (2). A negative electrode 6b is formed at the apex on the opposite side to the positive electrode (6a), and an antireflection film (7) is formed on the surface side of the diffusion layer (3). |
1. A semiconductor device, comprising: a semiconductor element, which has a flat surface formed by removing an apex part of a substantially spherical semiconductor crystal made of a p-type or n-type semiconductor; a diffusion layer or semiconductor thin-film deposition layer formed on a surface of the semiconductor element excluding the flat surface, and a substantially spherical pn junction formed via the diffusion layer or semiconductor thin-film deposition layer; and first and second electrodes, which are provided on the flat surface and at an apex on the opposite side to the flat surface respectively so as to face each other with a center of the semiconductor element interposed therebetween, and which are connected to both ends of the pn junction. 2. The semiconductor device according to claim 1, wherein the substantially spherical semiconductor crystal is fabricated by causing a semiconductor melt to solidify with the substantially spherical shape thereof maintained by utilizing a surface tension of the semiconductor melt. 3. The semiconductor device according to claim 1 or 2, wherein, when a protrusion exists near the surface of the semiconductor crystal, the flat surface of the semiconductor element is formed by removing the protrusion. 4. The semiconductor device according to claims 1 or 2, wherein a transparent insulating antireflection film is formed on the substantially spherical surface of the diffusion layer. 5. The semiconductor device according to claims 1 or 2, wherein the p-type or n-type semiconductor constituting the semiconductor crystal is a compound semiconductor selected from gallium arsenide (GaAs), indium phosphide (InP), gallium phosphide (GaP), gallium nitride (GaN), indium copper selenide (InCuSe), and silicon carbide (SiC). 6. The semiconductor device according to claims 1 or 2, wherein the semiconductor device is a light emitting device. 7. The semiconductor device according to claims 1 or 2, wherein the semiconductor device is a solar cell. 8. The semiconductor device according to claims 1 or 2, wherein the semiconductor device is a photodiode. 9. The semiconductor device according to claims 1 or 2, wherein the semiconductor device is a phototransistor. 10. A semiconductor device making method, comprising: a first step of making a substantially spherical semiconductor crystal made of a p-type or n-type semiconductor; a second step of producing a semiconductor element which has a flat surface formed by removing an apex part of the semiconductor crystal; a third step of forming a diffusion layer or semiconductor thin-film deposition layer on a surface of the semiconductor element excluding the flat surface and forming a substantially spherical pn junction via the diffusion layer or semiconductor thin-film deposition layer; and a fourth step of forming first and second electrodes, which are connected to both ends of the pn junction on the flat surface and at the apex on the opposite side to the flat surface respectively so as to face each other with a center of the semiconductor element interposed therebetween. 11. The semiconductor device fabrication method according to claim 10, wherein: in the third step, an insulating antireflection film is formed on the substantially spherical surface of the diffusion layer when the pn junction is formed; and, in the fourth step, electrode materials are provided on the flat surface and at the apex on the opposite side to the flat surface respectively so as to face each other with the center of the semiconductor element interposed therebetween, and first and second electrodes are formed from this pair of electrode materials. |
<SOH> BACKGROUND ART <EOH>Conventionally, research has been directed toward a technology that involves forming a pn junction, via a diffusion layer, on a surface of a small-diameter spherical semiconductor element made of a p-type or n-type semiconductor and then connecting a plurality of these spherical semiconductor elements in parallel to a common electrode, this technology being put to practical use for solar cells, semiconductor photocatalysts, and so forth. U.S. Pat. No. 3,998,659 discloses an example in which a solar cell is constituted by forming a p-type diffusion layer on the surface of a n-type spherical semiconductor, connecting the respective diffusion layers of a plurality of spherical semiconductors to a common film-like electrode (positive electrode), and then connecting the n-type core sections of these spherical semiconductors to a common film-like electrode (negative electrode). U.S. Pat. No. 4,021,323 discloses a solar energy converter (semiconductor module) having the following constitution. Plural p-type spherical semiconductor elements and plural n-type spherical semiconductor elements are placed in series, and connected to a common film-like electrode, and respective diffusion layers of these semiconductor elements are made into contact with a common electrolytic solution, and then by irradiating with solar light, electrolysis of the electrolytic solution is induced. So too in the case of the modules having spherical cells appearing in U.S. Pat. Nos. 4,582,588 and 5,469,020, because the spherical cells are attached by being connected to a sheet-like common electrode, a plurality of spherical cells are suitable for connecting in parallel. However, they are not suitable for serial connection. On the other hand, as shown in International Patent Publication Nos. WO98/15983 and WO99/10935, the inventor of the present invention has proposed a granular light emitting or light receiving semiconductor device in which a diffusion layer, pn junction, and a pair of electrodes are formed on a spherical semiconductor element made of a p-type semiconductor or an n-type semiconductor. Also, proposed is a semiconductor module, which is produced by connecting a plurality of the semiconductor device in series and then connecting a plurality of the serially connected bodies in parallel, and which can be applied to a solar cell, a photocatalyst device for electrolysis of water and so forth, a variety of light emitting devices, and color displays, and the like. In the case of this semiconductor module, when any semiconductor device of any serially connected body enters an open state due to failure, current no longer flows to the serial circuit including above failed semiconductor element, and the remaining normal semiconductor devices in the serially connected body also enter a breakdown state, whereby dropping of the output of the semiconductor module is generated. In addition, in the case of the spherical semiconductor devices having the positive and negative electrodes that were proposed by the present inventor, handling is a problem because the device is prone to rolling, and it is not easy to determine the position for forming the positive and negative electrodes nor to distinguish the positive and negative electrodes during assembly. Therefore, the inventor of this application undertook research with respect to a technology for forming a pair of flat surfaces on a spherical semiconductor element and then for forming electrodes on these flat surfaces. However, not only was there then a large number of processes for the electrode formation, it also became evident that it was still not easy to distinguish between the positive and negative electrodes and that this technology was not very advantageous in terms of mass producing the semiconductor module by using a multiplicity of spherical semiconductor devices. An object of the present invention is accordingly to provide a substantially spherical semiconductor device having one flat surface, is not prone to rolling and can be handled easily. Another object of the present invention is to provide a semiconductor device in which a first electrode is formed on the flat surface and a second electrode is formed at the apex on the opposite side to this electrode such that the center of the semiconductor device is interposed between the first and second electrodes, and in which the positive and negative electrodes can be easily distinguished. A further object of the present invention is to provide a making method for this semiconductor device. |
<SOH> BRIEF DESCRIPTION OF THE DRAWINGS <EOH>FIGS. 1 to 38 show embodiments of the present invention. FIGS. 1 ( a ) and 1 ( b ) are cross-sectional views of a spherical semiconductor crystal and a substantially spherical semiconductor crystal respectively; FIG. 2 is a cross-sectional view of a semiconductor element having a flat surface; FIG. 3 is a cross-sectional view of a semiconductor element having a diffusion-mask thin film; FIG. 4 is a cross-sectional view of the semiconductor element in FIG. 3 , an acid-resistant sheet and an acid-resistant wax; FIG. 5 is a cross-sectional view of a semiconductor element on which a partial diffusion-mask thin film remains; FIG. 6 is a cross-sectional view of a semiconductor element having a diffusion layer, a pn junction and an antireflection film; FIG. 7 is a cross-sectional view of a semiconductor element with pasting an electrode-forming aluminum paste and silver paste to the semiconductor element of FIG. 6 ; FIG. 8 is a cross-sectional view of a semiconductor element having a pair of electrodes formed by heat-treating the semiconductor element of FIG. 7 . FIG. 9 is a plan view of a lead frame; FIG. 10 is a cross-sectional view of the lowest lead frame and paste; FIG. 11 is a cross-sectional view of an intermediate lead frame and paste; FIG. 12 is a plan view of an assembly body produced by integrating a plurality of semiconductor devices and a plurality of lead frames; FIG. 13 is a front view of the assembly body; FIG. 14 is a plan view of a lead frame, and three sets of semiconductor modules molded with a light transmitting member made of a transparent synthetic resin; FIG. 15 is a cross-sectional view along the line XV-XV in FIG. 14 ; FIG. 16 is a plan view of a semiconductor module; FIG. 17 is a front view of a semiconductor module; and FIG. 18 is an equivalent circuit of the semiconductor module. FIG. 19 is a plan view of a lead frame and a one-set semiconductor module molded with a light transmitting member made of a transparent synthetic resin, relating to a second modified embodiment; FIG. 20 is a cross-sectional view along the line XX-XX in FIG. 19 ; FIG. 21 is a plan view of a semiconductor module relating to a third modified embodiment; and FIG. 22 is a cross-sectional view along the line XXII-XXII in FIG. 21 . FIG. 23 is a plan view of a base sheet relating to a fourth modified embodiment; FIG. 24 is a plan view of the base sheet with connecting leads; FIG. 25 is a plan view of a base sheet on which the semiconductor devices are mounted; FIG. 26 shows an end face view of an assembly body produced by assembling the base sheet and semiconductor devices; FIG. 27 shows an end face view of a semiconductor module comprising the base sheet, semiconductor devices and a light transmitting member; FIG. 28 shows the end face view of the semiconductor module in which the semiconductor module in FIG. 27 is partially modified; FIG. 29 is a plan view of a base sheet different from the one mentioned above; and FIG. 30 is a vertical cross-sectional view of the base sheet in FIG. 29 . FIG. 31 is a cross-sectional view of a semiconductor element relating to a fifth modified embodiment; FIG. 32 is a cross-sectional view of a semiconductor element having a silicon growth layer and a pn junction on the semiconductor element in FIG. 31 ; FIG. 33 is a cross-sectional view of a semiconductor element having an antireflection film on the semiconductor element in FIG. 32 ; and FIG. 34 is a cross-sectional view of a semiconductor device having positive and negative electrodes on the semiconductor element in FIG. 33 . FIG. 35 is a cross-sectional view of a semiconductor element relating to a sixth modified embodiment; FIG. 36 is a cross-sectional view of a semiconductor element having a p-type base layer on the semiconductor element in FIG. 35 ; FIG. 37 is a cross-sectional view of a semiconductor element having an n-type emitter layer on the semiconductor element in FIG. 36 ; and FIG. 38 is a cross-sectional view of an npn phototransistor. detailed-description description="Detailed Description" end="lead"? |
Method and system for node failure detection |
A distributed computer system, including a group of nodes. Each of the nodes have a network operating system, enabling one-to-one messages and one-to-several messages between said nodes, a first function capable of marking a pending message as in error, a node failure storage function, and a second function. The second function being responsive to the node failure storage function indicating a given node as failing, for calling said first function to force marking selected messages to that given node into error, the selected messages include pending messages which satisfy a given condition. |
1. A distributed computer system, comprising a group of nodes, each having: a network operating system enabling one-to-one messages and one-to-several messages between said nodes, a first function capable of marking a pending message as in error, a node failure storage function, and a second function, responsive to the node failure storage function indicating a given node as failing, for calling said first function to force marking selected messages to that given node into error, the selected messages comprising pending messages which satisfy a given condition. 2. The distributed computer system of claim 1, wherein the second function, responsive to the node failure storage function indicating a given node as failing, is adapted to call said first function to further force marking selected future messages to said given node into error, the selected messages comprising future messages which satisfy a given condition. 3. The distributed computer system of claim 1, wherein the node failure storage function is arranged for storing identifications of failing nodes from successive lack of response of such a node to an acknowledgment-requiring message. 4. The distributed computer system of claim 1, wherein the given condition comprises the fact a message specifies the address of said given node as a destination address. 5. The distributed computer system of claim 1, wherein: said group of nodes has a master node, said master node having a node failure detection function, capable of: repetitively sending an acknowledgment-requiring message from the master node to at least some of the other nodes, responsive to a given node failure condition, involving possible successive lack of responses from the same node, storing identification of that node as a failing node in the node failure storage function of the master node, and sending a corresponding node status update message to all other nodes in the group, and each of the non master nodes having a node failure registration function responsive to receipt of such a node status update message for updating a node storage function of the non master node. 6. The distributed computer system of claim 1, wherein the first and second functions are part of the operating system. 7. The distributed computer system of claim 5, wherein each node of the group having a node storage function for storing identifications of each node of the group and, responsive to the node failure storage function, updating identifications of failing nodes. 8. The distributed computer system of claim 1, wherein each node uses a messaging function called Transmission Transport Protocol. 9. The distributed computer system of claim 5, wherein the node failure detection function in master node uses a messaging function called User Datagram Protocol. 10. The distributed computer system of claim 5, wherein the node failure registration function in non master node uses a messaging function called User Datagram Protocol. 11. A method of managing a distributed computer system, comprising a group of nodes, said method comprising the steps of: detecting at least one failing node in the group of nodes, issuing identification of that given failing node to all nodes in the group of nodes, responsive to the step of issuing identification of that given failing node to all nodes in the group of nodes: storing an identification of that given failing node in at least one of the nodes, calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising pending messages which satisfy a given condition. 12. The method of claim 11, wherein the step of calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising pending messages which satisfy a given condition further comprises calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising future messages which satisfy a given condition. 13. The method of claim 11, wherein the method further comprises repeating in time the steps of: detecting at least one failing node in the group of nodes, issuing identification of that given failing node to all nodes in the group of nodes, responsive to the step of issuing identification of that given failing node to all nodes in the group of nodes: storing an identification of that given failing node in at least one of the nodes, calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising pending messages which satisfy a given condition. 14. The method of claim 11, wherein the given condition in the step of calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error comprises the fact a message specifies the address of said given node as a destination address. 15. The method of claim 11, wherein the step of detecting at least one failing node in the group of nodes further comprises: electing one of the nodes as a master node in the group of nodes, repetitively sending an acknowledgment-requiring message from a master node to all nodes in the group of nodes, responsive to a given node failure condition, involving possible successive lack of responses from the same node, storing identification of that node as a failing node in the master node. 16. The method of claim 15, wherein of detecting at least one failing node in the group of nodes further comprises storing identification of the given failing node in a master node list. 17. The method of claim 16, wherein the step of detecting at least one failing node in the group of nodes further comprises deleting identification of the given failing node in the master node list. 18. The method of claim 11, wherein the step of issuing identification of that given failing node to all nodes in the group of nodes further comprises sending the master node list to all nodes in the group of nodes. 19. The method of claim 11, wherein the step of storing an identification of that given failing node in at least one of the nodes further comprises updating a node list in all nodes with the identification of the given failing node. 20. The method of claim 11, wherein the step of calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising pending messages which satisfy a given condition, further comprises calling the function in a network operating system of at least one node. 21. A software product, comprising the software functions used in a distributed computer system, comprising a group of nodes, each having: a network operating system enabling one-to-one messages and one-to-several messages between said nodes, a first function capable of marking a pending message as in error, a node failure storage function, and a second function responsive to the node failure storage function indicating a given node as failing, for calling said first function to force marking selected messages to that given node into error, the selected messages comprising pending messages which satisfy a given condition. 22. A software product, comprising the software functions for use in a method of managing a distributed computer system, comprising a group of nodes, said method comprising the steps of: detecting at least one failing node in the group of nodes, issuing identification of that given failing node to all nodes in the group of nodes, responsive to the step of issuing identification of that given failing node to all nodes in the group of nodes: storing an identification of that given failing node in at least one of the nodes, calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising pending messages which satisfy a given condition. 23. A network operating system, comprising a software product, comprising the software functions used in a distributed computer system, comprising a group of nodes, each having: a network operating system enabling one-to-one messages and one-to-several messages between said nodes, a first function capable of marking a pending message as in error, a node failure storage function, and a second function, responsive to the node failure storage function indicating a given node as failing, for calling said first function to force marking selected messages to that given node into error, the selected messages comprising pending messages which satisfy a given condition. 24. A network operating system, comprising a software product comprising the software functions for use in a method of managing a distributed computer system, comprising a group of nodes, said method comprising the steps of: detecting at least one failing node in the group of nodes, issuing identification of that given failing node to all nodes in the group of nodes, responsive to the step of issuing identification of that given failing node to all nodes in the group of nodes: storing an identification of that given failing node in at least one of the nodes, calling a function in at least one of the nodes to force marking selected messages between that given failing node and said node into error, the selected messages comprising pending messages which satisfy a given condition. |
Peptides conjugates, their derivatives with metal complexes and use thereof for magnetic resonance imaging (mri) |
The present invention relates to a novel class of contrast agents for use in nuclear magnetic resonance (MRI), to identify and locate primary human tumours and their metastases which over-express type CCK A and/or type B cholecystokinin receptors, and/or type SSTR 1-5 somatostatin receptors. |
1. Compounds of general formula (I): [A]t[P]—[B]v (I) wherein: B represents one or more straight, branched or cyclic peptides of general formula (II) or (III) (AA0)w-AA1-AA2-AA3-Gly-Trp-AA6-Asp-AA8-R2 (II) wherein: AA0 is any amino acid in L or D configuration; AA1 is Asp or Glu; AA2 is Tyr or SO3H-Tyr; AA3 is Met, Me or Leu; AA6 is Met, Me or Leu; AA8 is Phe or the corresponding amino alcohol, AA′1, AA′3 AA′6 and AA′8 are any amino acid, either natural or not, in L or D configuration; AA′8 can also be an amino alcohol derivative from any amino acid, either natural or not, in L or D configuration; R2 is a hydroxy, amino or C1-C4 alkoxy group; v is an integer of 1 to 5; w is zero or 1; P represents a straight or branched polymeric chain able to covalently bind t A units and bonded by covalent bonds to one or more units of B; t is an integer ranging from 2 to 100; A represents a cyclic or acyclic chelating agent covalently bound to P and containing 6 to 8 coordination sites selected from amino, pyridino, carboxy, phosphonic, phosphinic, hydroxyl and carboxamido groups, said groups A being able to chelate bi- trivalent metals having atomic numbers ranging from 21 to 29, 42, 44 or form 57 to 71. 2. Compounds as claimed in claim 1 in the form of complexes with the bi-trivalent ions of metal elements having atomic numbers ranging from 21 to 29, 42, 44 or from 57 to 71, with paramagnetic metals, in particular with Fe, 10 Cu, Cr, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Mn, as well as the salts thereof with physiologically compatible bases or acids. 3. Compounds as claimed in claim 2 in the form of complexes with the bi-trivalent ions of Fe(2+), Fe(3+), Cu(2+), Cr(2+), Cr(3+), Eu (3+), Gd(3+), Tb(3+), Dy(3+), Ho(3+), Er(3+), Yb(3+), Mn(2+) and Mn(3+) as well as the salts thereof with physiologically compatible bases or acids. 4. Compounds as claimed in claim 3 in the form of complexes obtained with the bi-trivalent ions of Eu(3+), Gd(3+), Dy(3+), Mn(2+) and Mn(3+), as well as the salts thereof with physiologically compatible bases or acids. 5. Compounds as claimed in any one of the above claims, wherein the peptide residues B of formula (It) and (III), are selected from: Gly-Glu-Tyr-Met-Gly-Trp-Met-Asp-Phe-NH2 Gly-Asp-Tyr-Met-Gly-Trp-Leu-Asp-Phe-OH Gly-Glu-Tyr(SO3H)-Met-Gly-Trp-Leu-Asp-Phe-NH2 6. Compounds as claimed in any one of claims 1 to 5 wherein the chelating group A is selected from the group consisting of: residues of polyaminopolycarboxylic acids and derivatives thereof, polyaminophosphonic acids and derivatives thereof, polyaminophosphinic acids and derivatives thereof, texaphyrines, porphyrins, phthalocyanines. 7. Compounds as claimed in claim 6 wherein the chelating group A is selected from the group consisting of: ethylenediaminotetraacetic acid (EDTA), diethylenetriaminopentaacetic acid (DTPA), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (DO3A), [10-(2-hydroxypropyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (HPDO3A), 4-carboxy-5,8,11-tris(carboxymethyl)-1-phenyl-2-oxa-5,8,11-triazatridecan-13-oic acid (BOPTA), N-[2-[bis(carboxymethyl)amino]-3-(4-ethoxyphenyl)propyl]-N-[2-[bis(carboxymethyl)amino]ethylglycine (EOB-DTPA), N,N-bis[2-(carboxymethyl)[(methylcarbamoyl)methyl]amino]ethyl]-glycine (DTPA-BMA), 2-methyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (MCTA), (α,α′,α″,α′″)-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetracetic acid (DOTMA); the residue of a polyaminophosphonic acid ligand and derivatives thereof, polyaminophosphinic acid and derivatives thereof, in particular ethylenedinitrilotetrakis(methylphosphonic) acid (EDTP); 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis[methylene(methylphosphonic)]acid and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis[methylene-(methylphosphinic)]acid. 8. Compounds as claimed in any one of claims 1 to 7, wherein A is selected from: 9. Compounds as claimed in any one of claims 1 to 8 wherein the polymer P compound is a polipeptide, a polyamine, polyethyleneiminopolyacetic acid ester derivatives, poly(alkyleneoxides), alpha(polyamino)poly-(alkyleneoxides), poly(ethyleneoxide) (PEO), poly(propyleneoxide) (PPO), poly(butyleneoxide), polyoxypropylene glycol, polyoxyethylene glycol, polyoxyalkylene glycol, polyalkyl esters, polyalkylcyanoacrylates, polymethylcyanoacrylates, polyethylcyanoacrylates, polybutylcyanoacrylates, polysobutylcyanoacrylates and copolymers thereof. 10. Compounds as claimed in claim 9 wherein the polymer compound P is a polylysine or a poly-L-lysine or a Lys-β-Ala or Dap-β-Ala copolymer. 11. Compounds as claimed in any one of the above claims selected from: (DTPA-GLU)4(Lys)2Lys-Gly-CCK8 (DTPA-GLU)3(Lys)2Lys-Gly-CCK8 (DTPA-GLU)4(Lys)2Lys-Gly-Vapreotide (DO3A-Ar)4(Lys)2Lys-Gly-Vapreotide (Lys(DTPA-GLU)-βAla)4Lys(DTPA-GLU)-Gly-CCK8 (Lys(DTPA-GLU)-βAla)9Lys(DTPA-GLU)-Gly-CCK8 (Lys(DTPA-GLU)-βAla)4Lys(DTPA-GLU)-Gly-Vapreotide (Lys(DTPA-GLU)-βAla)9Lys(DTPA-GLU)-Gly-Vapreotide (Dap(DTPA-GLU)-βAla)4-Dap(DTPA-GLU)-Gly-CCK8 (Dap(DTPA-GLU)-βAla)9-Dap(DTPA-GLU)-Gly-CCK8 (Dap(DTPA-GLU)-βAla)4-Dap(DTPA-GLU)-Vapreotide (Dap(DTPA-GLU)-βAla)9-Dap(DTPA-GLU)-Vapreotide 12. Pharmaceutical and diagnostic composition containing a compound of claims 1-11 in admixture with a suitable carrier. 13. The use of the compounds of claims 1 to 11 and of the salts thereof for the preparation of diagnostic formulations used in MRI investigations, for imaging and recording images of organs and/or tissues. 14. The use as claimed in claim 13, for in vitro and/or in vivo imaging and recording images of cells, tissues or organs in which tumours or primary tumour pathologies or metastases are present. 15. Compounds of formula: A′-Dap-Fmoc wherein A′ is a unit of formula A as defined above having the carboxy or phosphonic moieties suitably protected, Dap is a diamino-acid, specifically 2,3-diaminopropionic acid and Fmoc is (9H-fluoren-9-ylmethoxy)carbonyl. 16. A compound according to claim 15, which is 3-[[(4S)-4-[bis[2-[bis[2-(1,1-dimethylethoxy)-2-oxoethyl]amino]ethyl]amino]-5-(1,1-dimethylethoxy)-1,5-dioxopentyl]amino]-N-[(9H-fluoren-9-ylmethoxy)carbonyl]-L-alanine. |
Prime ring substituted thyroid receptor antagonists for the treatment of cardiac and metabolic disorders |
This invention relates to novel compounds which are thyroid receptor ligands, preferably antagonists, and to methods for using such compounds in the treatment of cardiac and metabolic disorders, such as cardiac arrhythmias, thyrotoxicosis, subclinical hyperthyrodism and liver diseases. |
1. A compound according to the general formula: or a pharmaceutically acceptable salt thereof, wherein: R1 is independently selected from: carboxylic acid (—CO2H); carboxylic acid ester (—CO2Rc); alpha-hydroxy carboxylic acid (—CH(OH)CO2H); alpha-amino carboxylic acid (—CH(NH2)CO2H); phosphonic acid (—PO(OH)2); phosphamic acid (—PO(OH)NH2); sulphonic acid (—SO2OH); hydroxamic acid (—CONHOH); oxamic acid (—NHCOCO2H); and malonamic acid (—NHCOCH2CO2H), and bioisosteric equivalents of any thereof; R2 and R3 are the same or different and independently selected from: chlorine; bromine; iodine; C1-4 alkyl or haloalkyl; C2-4 alkenyl or haloalkenyl; C2-4 alkynyl or haloalkynyl, or a bioisosteric equivalent of any thereof; R4 is selected from: halogen; C1-4 alkyl; C2-4 alkenyl; and C2-4 alkynyl, each being optionally substituted with 1, 2 or 3 halogen atoms, which may be the same or different, or a bioisosteric equivalent of any thereof; X is selected from: —CH2—CH2—; —CH2—CH2—CH2; —CH2═CH2—; —CH2═CH—CH2—; —C═C—; and —C≡C—CH2; R5 is selected from C6-10 aryl; C1-9 heteroaryl; and C5-10 cycloalkyl, each being optionally substituted with 1, 2, or 3 groups of Rb which may be the same or different; Rb is selected from: halogen; —CN; —CO2H; —CHO; —NO2; C1-4 alkyl; C2-4 aklenyl; C2-4 alkynyl; C1-4 alkoxy; C2-4 alkenoxy; C2-4 alkynoxy; C1-4 alkylthio; C2-4 alkenylthio; C2-4 alkynylthio; —(CH2)n—OH; —(CH2)n—O(C1-4); —(CH2)n—NH2; —(CH2)n—NH(C1-4); and —(CH2)n—N(C1-4)2, or a bioisosteric equivalent of any thereof; Rc is selected from C1-4 alkyl; C2-4 alkenyl; and C2-4 alkynyl; n is 1 or 2; and stereoisomers; prodrug ester forms, and radioactive forms thereof. 2. A compound according to claim 1 wherein R1 is selected from: carboxylic acid (—CO2H); alpha-hydroxy carboxylic acid (—CH(OH)CO2H); and alpha-amino carboxylic acid (—CH(NH2)CO2H). 3. A compound according to claim 1 wherein R3 is bromine. 4. A compound according to claim 1 wherein R4 is isopropyl. 5. A compound according to claim 1 wherein R5 is C6 aryl or C1-5 heteroaryl. 6. A compound according to claim 1 wherein X is —C═C—. 7. A compound according to claim 1 wherein R1 is a carboxylic acid (—CO2H) and X is —C═C—. 8. A compound according to claim 1 wherein R1 is a carboxylic acid (—CO2H); R2 and R3 is bromine; R4 is isopropyl; R5 is C6 aryl or C1-5 heteroaryl. 9. A compound according to claim 1 which is: 3,5-dibromo-4-[4-hydroxy-3-isopropyl-5-((E)-styryl)phenoxy]benzoic acid (E1); 3-{3,5-dibromo-4-[4-hydroxy-3-isopropyl-5((E)-styryl)phenoxy]phenylpropionic acid (E2); 3-{3,5-dibromo-4-[4-hydroxy-3-isopropyl-5((E)-2-pyridin-4-yl-vinyl)phenoxy]-phenyl propionic acid (E3); 3-{3,5-dibromo-4-[4-hydroxy-3-isopropyl-5((E)-2-pyridin-2-yl-vinyl)phenoxy]phenyl}-propionic acid (E4); 3-{3,5-dibromo-4-[4-hydroxy-3-isopropyl-5-((E)-2-pyrazine-2-yl-vinyl)phenoxy]phenyl}-propionic acid (E5); 3-(3,5-dibromo-4-{3-[(E)-2-(4-dimethylaminomethylphenyl)vinyl]-4-hydroxy-5-isopropyl phenoxy}phenyl)propionic acid (E6); 3-(3,5-Dibromo-4-{4-hydroxy-3-isopropyl-5-[(E)-2-(4-methylthiazol-5-yl)vinyl]phenoxy}-phenyl)propionic acid (E7); 4-((E)-2-{5-[2,6-Dibromo-4-(2-carboxyethyl)phenoxy]-2-hydroxy-3-isopropylphenylvinyl)-benzoic acid (E8); 3-{3,5-Dibromo-4-[4-hydroxy-3-isopropyl-5-(2-pyridin-4-yl-ethyl)-phenoxy]-phenyl}propionic acid (E9); 3-[3,5-Dibromo-4-(4-hydroxy-3-isopropyl-5-phenetyl-phenoxy)-phenyl]-propionic acid (E10); 3-[3,5-Dibromo-4-[4-hydroxy-3-isopropyl-5 (E)-styryl-phenoxy)phenyl]-2-hydroxy propionic acid (E11); 3-{3,5-Dibromo-4-[4-hydroxy-3-isopropyl-5-((E)-2-pyridin-4-yl-vinyl)-phenoxy]-phenyl}-2-hydroxy propionic acid (E12); 3-[3,5-Dibromo-4-[4-hydroxy-3-isopropyl-5-phenylethyl-phenoxy)-phenyl]-2-hydroxy-propionic acid (E13); 3-{3,5-Dibromo-4-[4-hydroxy-3-isopropyl-5-(2-pyridin-4-yl-ethyl)-phenoxy]-phenyl}-2-hydroxy propionic acid (E14); and pharmaceutically acceptable salts thereof and stereoisomers thereof. 10. A compound according to claim 1, which has one or more asymmetric centers and thus can exist in the form of racemates, single and multiple enantiomers, individual diastereomers, with all possible isomers, and mixtures thereof. 11. (canceled) 12. A pharmaceutical composition comprising an effective amount of a compound according to claim 1 or a pharmaceutically effective salt thereof, together with a pharmaceutically acceptable carrier. 13. A method for preventing, inhibiting or treating a disease dependent on the expression of a T3 regulated gene or associated with metabolic dysfunction, which method comprises administering to a patient in need of treatment a therapeutically effective amount of a compound according to claim 1. 14. The method according to claim 13 wherein the disease is cardiac arrhythmias, thyrotoxicosis, subclinical hyperthyroidism, skin disorders, or a liver disease. 15. The method according to claim 14 wherein the disease is a skin disorder or skin disease. 16. The method according to claim 15 wherein the skin disorder or skin disease is: keloids, lichen planus, ichtyosis, acne, psoriasis, Dernier's disease, eczema, atopic dermatitis, chloracne, pityriasis, or hirsuitism. 17. The method according to claim 14 wherein the disease is a liver disorder or liver disease. 18. The method according to claim 17 wherein the liver disorder or liver disease is: chronic alcoholism, acute hepatitis, chronic hepatitis, hepatitis C-induced liver cirrhosis, or liver fibrosis. 19. A method to treat skin disorders or diseases comprising the step of administering to a patient a pharmaceutical composition comprising the compound of claim 1 and a retinoid or a Vitamin D analog. 20-27. (canceled) |
<SOH> BACKGROUND OF THE INVENTION <EOH>Nuclear hormone receptors comprise a class of intracellular, mostly ligand-regulated transcription factors, which include receptors for thyroid hormones. Thyroid hormones exert profound effects on growth, development and homeostasis in mammals. They regulate important genes in intestinal, skeletal and cardiac muscles, liver and the central nervous system, and influence the overall metabolic rate, cholesterol and triglyceride levels, heart rate, and affect mood and overall sense of well being. There are two major subtypes of the thyroid hormone receptor, TRα and TRβ, expressed from two different genes. Differential RNA processing results in the formation of at least two isoforms from each gene. The TRα 1 TRβ 1 and TRβ2 isoforms bind thyroid hormone and act as ligand-regulated transcription factors. The TRα 2 isoform is prevalent in the pituitary and other parts of the central nervous system, does not bind thyroid hormones, and acts in many contexts as a transcriptional repressor. In adults, the TRβ 1 isoform is the most prevalent form in most tissues, especially in the liver and muscle. The TRα 1 isoform is also widely distributed, although its levels are generally lower than those of the TRP I isoform. A growing body of data suggest that many or most effects of thyroid hormones on the heart, and in particular on the heart rate and rhythm, are mediated through the TRβ 1 isoform, whereas most actions of the hormones on the liver, muscle and other tissues are mediated more through the β-forms of the receptor. It is believed that the α-isoform of the receptor is the major influence on heart rate for the following reasons: (i) tachycardia is very common in the syndrome of generalized resistance to thyroid hormone in which there are defective TRβ-isoforms, and consequently high circulating levels of T 4 and T 3 ; (ii) Tachycardia was observed in the only described patient with a double deletion of the TRβ gene (Talceda et al, J. Clin. Endrocrinol . & Metab. 1992, 74, 49); (iii) a double knockout TRα gene (but not β-gene) in mice showed bradycardia and lengthening of action potential compared to control mice (Functions of Thyroid Hormone Receptors in Mice: D. Forrest and B. Vennström, Thyroid, 2000, 10, 41-52); (iv) western blot analysis of human myocardial TRs show presence of the TRα 1 , TRα 2 and TRβ 2 proteins, but not TRβ 1 . If the indications above are correct, an a-selective thyroid hormone receptor antagonist that interacts selectively with the heart would offer an attractive alternative treatment of heart related disorders, such as atrial and ventricular arrhythmias. Atrial fibrillation (AF) is the most common type of sustained arrhythmia encountered in primary care practice and is significantly more common in elderly patients, thus reflecting a reduction in the threshold for AF with age. Pharmacological treatment of AF involves the following types of anti-arrhythmic drugs according to Vaughan-Williams classification: (i) of class I such as disopyramide and flecainide (sodium channel blockers); (ii) of class III such as amiodarone (potassium channel blocker, prolongation of repolarization); (iii) of class IV such as verapamil and dilitazem (calcium channel blockers). Many patients are also subjected to electric cardioversions in order to convert atrial fibrillation into sinus rhythm. It should be noted that current therapies are associated with pro-arrhythmic risks and anti-arrhythmic agents often have insufficient efficacy partly because effective doses are limited by side-effects. Ventricular arrhythmia, especially sustained ventricular tachycardia (VT) and ventricular fibrillation (VF), is the main cause of death associated with heart attack. Historically, three types of antiarrhythmic agents, class I agents, β-adrenergic blockers (class II), amiodarone and sotalol, appeared to offer the best scope for mortality reduction in patients with cardiac disease by preventing the occurrence of VT/VF. The outcome of CAST (Cardiac Arrhythmia Supression Trial, N. Engl. J. Med., 321 (1989) 406-412) and its successor SWORD (Survival With Oral D-sotatol trial, 1994) created much concern regarding the potential of class I agents and sotalol. It was found that class I agents did not decrease mortalities in patient groups at risk for sudden cardiac death. For some subsets of patients, class I agents even proved to increase mortality. The SWORD trial was stopped when sotalol proved to be associated with higher death rate in patients, than the placebo. A consequence of these results is that the use of implantable defibrillators and surgical ablation have increased and that the trend in the industry has been towards the development of highly specific class III agents. Some of these channel blockers have been withdrawn from clinical development due to proarrhythmic effects and the subject remains under intensive debate. In this context it should be noted that amiodarone, despite its complex pharmacokinetics, mode of action (amiodarone is not regarded as a pure class III agent) and numerous side effects, is currently considered by many to be the most effective agent in the control of both atrial and ventricular arrhythmia. Thyrotoxicosis is the clinical syndrome that results when tissues are exposed to elevated levels of circulating thyroid hormones, thyroxine (3,5,3′,5′-tetraiodo-L-thyronine, or T 4 ) and triiodothyronine (3,5,3′-triiodo-L-thyronine, or T 3 ). Clinically, this state often manifests itself in weight loss, hypermetabolism, lowering of serum LDL levels, cardiac arrhythmia, heart failure, muscle weakness, bone loss in postmenopausal women, and anxiety. In most instances, thyrotoxicosis is due to hyperthyroidism, a term reserved for disorders characterized by overproduction of thyroid hormones by the thyroid gland. The ideal treatment of hyperthyroidism would be the elimination of its cause. This is however not possible in the more common diseases producing thyroid hypersecretion. At present, treatment of hyperthyroidism is directed to reducing overproduction of thyroid hormones by inhibiting their synthesis or release, or by ablating thyroid tissue with surgery or radioiodine. Drugs inhibiting thyroid hormone synthesis, release, or peripheral conversion of T 4 to T 3 include antithyroid drugs (thionamides), iodide, iodinated contrast agents, potassium perchlorate and glucocorticoids. The main action of antithyroid drugs such as methimazole (MMI), carbimazole, and propylthiouracil (PTU), is to inhibit the organification of iodide and coupling of iodotyrosines, thus blocking the synthesis of thyroid hormones. As they neither inhibit iodide transport nor block the release of stored thyroid hormones, control of hyperthyroidism is not immediate and in most cases requires 2 to 6 weeks. Factors that determine the speed of restoration of euthyroidism include disease activity, initial levels of circulating thyroid hormones, and intrathyroidal hormone stores. Serious side effects are not common with antithyroid drugs. Agranulocytosis is the most feared problem and has been observed with both MMI or PTU treatment. Elderly persons may be more susceptible to this side effect, but agranulocytosis can occur in younger age groups, although less frequently. Inorganic iodide given in pharmacological doses (as Lugol's solution or as saturated solution of potassium iodide, SSKI) decreases its own transport into the thyroid, thus inhibiting iodide organification (the Wolff-Chaikoff effect), and rapidly blocks the release of T 4 and T 3 from the gland. However, after a few days or weeks, its antithyroid action is lost, and thyrotoxicosis recurs or may worsen. Short-term iodide therapy is used to prepare patients for surgery, usually in combination with a thionamide drug. Iodide is also used in the management of severe thyrotoxicosis (thyroid storm), because of its ability to inhibit thyroid hormone release acutely. Perchlorate interferes with accumulation of iodide by the thyroid. Gastric irritation and toxic reactions limit the long-term use of perchlorate in the management of hyperthyroidism. Glucocorticoids in high doses inhibit the peripheral conversion of T 4 to T 3 . In Graves' hyperthyroidism, glucocorticoids appear to decrease T 4 secretion by the thyroid, but the efficiency and duration of this effect is unknown. The aim of surgical treatment or radioiodine therapy of hyperthyroidism is to reduce the excessive secretion of thyroid hormones by removal or destruction of thyroid tissue. Subtotal or near-total thyroidectomy is performed in Graves' disease and toxic multinodular goiter. Restoration of euthyroidism before surgery is mandatory. The classical approach combines a course of thionamide treatment to restore and maintain euthyroidism, and the preoperative administration of iodide for approximately 10 days in order to induce involution of the gland. Propranolol and other beta-adrenergic antagonist drugs are useful in controlling tachycardia and other symptoms of sympathetic activation. A high affinity ThR antagonist would in principle have the ability to restore euthyrodism quicker than any of the above agents, considered that its action is competitive for the ThR receptor. Such an agent could be used either alone or in combination with the above drugs, or alternatively before an ablative treatment. It may also serve as a safer substitute for antithyroid drugs, especially in elderly patients at a high risk of agranulocytosis. Furthermore, hyperthyrodism can aggravate pre-existing heart disease and also lead to atrial fibrillation (AF), congestive heart failure, or worsening of angina pectoris. In the elderly patient, often with mild but prolonged elevation of plasma thyroid hormones, symptoms and signs of heart failure and complicating AF may dominate the clinical picture and mask the more classical endocrine manifestations of the disease. |
Printer |
A printer prints on two sides of a sheet of paper having continuous pages, and discharges the paper to a postprocessor for disconnecting the paper in page units and storing the disconnected sheets of paper in a stack. Depending on whether the postprocessor has an inversion mechanism that inverts front and back sides of the paper, sides of the paper on which data for the front side of the paper and data for the back side of the paper are printed are switched. With this arrangement, even when the postprocessor does not have the inversion mechanism, the front side data is printed on the back side, and the back side data is printed on the front side thereby enabling the postprocessor to stack the disconnected sheets of paper in page units in the order of page numbers. |
1. A printer which is capable of printing on a front side and a back side of a sheet of paper having continuous pages, and discharging the paper to a postprocessor for disconnecting the paper in page units and storing the disconnected sheets of paper in a stack, the printer comprising: a first printing unit that prints a first side of the paper; a second printing unit that prints a second side of the paper; a deciding unit that determines whether the postprocessor has an inversion mechanism that inverts front and back sides of the paper discharged from the printer; and a control unit that determines which one of the first and second printing units prints a first print data portion for the first side of the paper and a second print data portion for the second side of the paper in the print data respectively, based on presence or absence of the inversion mechanism. 2. A printer according to claim 1, wherein when two-side printing is instructed, and when the postprocessor does not have the inversion mechanism, the control unit supplies the first print data portion of the print data for the first side of the paper to the second printing unit such that this print data portion is to be printed on the second side of the paper, and supplies the second print data portion for the second side of the paper to the first printing unit such that this print data portion is to be printed on the first side of the paper, and when the postprocessor has the inversion mechanism, the control unit supplies the first print data portion to the first printing unit such that this print data portion is to be printed on the first side of the paper, and supplies the second print data portion to the second printing unit such that this print data portion is to be printed on the second side of the paper. 3. A printer according to claim 1, wherein when the postprocessor does not have the inversion mechanism, and when one-side printing is instructed, the control unit supplies the first print data portion to the second printing unit such that this print data portion is to be printed on the second side of the paper, and supplies white paper data to the first printing unit such that this data is to be printed on the first side of the paper, thereby enabling the first printing unit and the second printing unit to execute two-side printing. 4. A printer according to claim 1, wherein the deciding unit determines presence or absence of the inversion mechanism based on flag information that is set by a predetermined setting unit and that indicates whether the postprocessor has the inversion mechanism. 5. A printer which is capable of printing on a first side and a second side of a sheet of paper having continuous pages, the printer comprising: a first printing unit that prints on the first side of the paper; a second printing unit that prints on the second side of the paper; a postprocessing unit that disconnects a printed sheet of paper in page units, and stores the disconnected sheets of paper in a stack; a deciding unit that determines whether the postprocessing unit has an inversion mechanism that inverts front and back sides of the printed sheets of paper; and a control unit that determines which one of the first and second printing units prints a first print data portion for the first side of the paper and a second print data portion for the second side of the paper in the print data respectively, based on presence or absence of the inversion mechanism. 6. A printer according to claim 2, wherein when the postprocessor does not have the inversion mechanism, and when one-side printing is instructed, the control unit supplies the first print data portion to the second printing unit such that this print data portion is to be printed on the second side of the paper, and supplies white paper data to the first printing unit such that this data is to be printed on the first side of the paper, thereby enabling the first printing unit and the second printing unit to execute two-side printing. 7. A printer according to claim 3, wherein the deciding unit determines presence or absence of the inversion mechanism based on flag information that is set by a predetermined setting unit and that indicates whether the postprocessor has the inversion mechanism. |
<SOH> BACKGROUND OF THE INVENTION <EOH>1. Field of the Invention The present invention relates to a printer that discharges a sheet of paper having continuous pages to a postprocessor that disconnects this paper into sheets of paper in page units, and stores the disconnected sheets of paper in a stack. Particularly, the invention relates to a printer that switches print sides of the sheets based on presence or absence of an inversion mechanism that inverts the front and back sides of the sheets in the postprocessor. 2. Description of the Related Art There are printers that can print on both sides of a sheet of paper having continues pages. The sheet of paper printed by the printer is stored into a body stacker that is incorporated in the printer main body or into a stacker of a postprocessor (for example, a Burster Trimmer Stacker) (hereinafter referred to as a “BTS stacker”) that receives the sheet of paper discharged from the printer, disconnects the printed paper into sheets of paper in page units by cutting or by bursting, and stores the disconnected sheets of paper in a stack. FIG. 7 is a view showing the storage of paper. A printer 1 has a lever 2 that selects a stacker which stores a printed sheet of paper. FIG. 7A illustrates that the lever 2 selects a body stacker 3 that is incorporated in the printer 1 . The printed sheet of paper is folded in page units, and is stored into the body stacker 3 . On the other hand, FIG. 7B illustrates that the lever 2 selects a BTS stacker 6 of a postprocessor 5 . The sheet or paper printed by the printer 1 enters the postprocessor 5 from a vent 4 of the printer. A disconnection mechanism not shown disconnects the sheet of paper into sheets of paper in page units, and stores the disconnected paper in a stack in the BTS stacker 6 . When sheets of paper printed on both sides of the paper are stored into the BTS stacker 6 of the postprocessor 5 , the following problems arise. FIG. 8 is a view showing a page layout of a two-side printed sheet of paper that is discharged from the printer. A first front side of the printed sheet of paper is the first page, and a back side of this paper is the second page. A front side that continues after the first page is the third page. A back side that continues after the second page is the fourth page. In other words, odd pages appear on the front side, and even pages appear on the back side of the paper. Therefore, in this page layout, the printed sheet of paper enters the postprocessor 5 , and is disconnected into sheets of paper in page units. As shown in FIG. 8A , the disconnected sheets of paper are stacked on the BTS stacker 6 , with page numbers appearing in the order of 2, 1, 4, 3, 6, 5, and so on from the bottom. Consequently, the page numbers are not continuous. On the other hand, some postprocessor 5 has an inversion mechanism that inverts the front and back sides of the sheets of paper that are discharged from the printer. When the inversion mechanism is used, the page layout is reverse of the above, with the front side being an even page and the back side being an odd page, as shown in FIG. 8B . Therefore, when the sheet of paper is disconnected into sheets of paper in page units, and the disconnected sheets of paper are stacked on the BTS stacker 6 , the page numbers appear as 1, 2, 3, 4, 5, and so on in this order from the bottom. Consequently, the page numbers are continuous in good order. As explained above, when the postprocessor 5 has the inversion mechanism, the printed sheets of paper are stored in the BTS stacker 6 in a state in which the page numbers are continuous. However, when the postprocessor 5 does not have the inversion mechanism (that is, when the printed sheet of paper enters the postprocessor 5 in a state of being discharged from the printer 1 , and when the paper disconnected into sheets of paper in page units are stored), the printed sheets of paper are stacked on the BTS stacker 6 in a state in which the page numbers are not continuous. |
<SOH> SUMMARY OF THE INVENTION <EOH>It is an object of the present invention to provide a printer that discharges a printed sheet of paper to a postprocessor for disconnecting the paper into sheets of paper in page units and storing the disconnected sheets of paper in a stack, wherein the postprocessor can stack the printed sheets of paper on a stacker of the postprocessor such that page numbers are in a continuous order regardless of presence or absence of an inversion mechanism that inverts the front and back sides of the sheets. In order to achieve the above object, according to one aspect of the invention, there is provided a printer which is capable of printing on a front side and a back side of a sheet of paper having continuous pages, and discharging the paper to a postprocessor for disconnecting the paper in page units and storing the disconnected sheets of paper in a stack, the printer comprising: a first printing unit that prints a first side of the paper; a second printing unit that prints a second side of the paper; a deciding unit that determines whether the postprocessor has an inversion mechanism that inverts front and back sides of the paper that is discharged from the printer; and a control unit that determines which one of the first and second printing units prints a first print data portion for the first side of the paper and a second print data portion for the second side of the paper in the print data respectively, based on presence or absence of the inversion mechanism. Specifically, when two-side printing is instructed, and when the postprocessor does not have the inversion mechanism, the control unit supplies the first print data portion of the print data for the first side of the paper to the second printing unit such that this print data portion is to be printed on the second side of the paper, and supplies the second print data portion for the second side of the paper to the first printing unit such that this print data portion is to be printed on the first side of the paper. When the postprocessor has the inversion mechanism, the control unit supplies the first print data portion to the first printing unit such that this print data portion is to be printed on the first side of the paper, and supplies the second print data portion to the second printing unit such that this print data portion is to be printed on the second side of the paper. As explained above, when the postprocessor that is connected to the printer does not have the inversion mechanism, the print data for the first side (i.e., the first print data portion) is printed on the second side of the paper, and the print data for the second side (i.e., the second print data portion) is printed on the first side of the paper. With this arrangement, the paper that is disconnected into page units by the postprocessor can be stacked in the order of page numbers. Further, when one-side printing is instructed, and when the postprocessor does not have the inversion mechanism, the control unit supplies the first print data portion to the second printing unit such that this print data portion is to be printed on the second side of the paper, and supplies white paper data to the first printing unit such that this data is to be printed on the first side of the paper thereby enabling the first printing unit and the second printing unit to execute two-side printing. Accordingly, in one-side printing, even when the postprocessor does not have the inversion mechanism, when the print side of each sheet of paper stacked on the stacker of the postprocessor is faced upward, the sheets of paper can be stacked in correct order of pages starting from the first page. Further, the deciding unit determines the presence or absence of the inversion mechanism based on flag information that is set by a predetermined setting unit and that indicates whether the postprocessor has the inversion mechanism. In order to achieve the above object, according to another aspect of the invention, there is provided a printer which is capable of printing on a first side and a second side of a sheet of paper having continuous pages, the printer comprising: a first printing unit that prints on the first side of the paper; a second printing unit that prints on the second side of the paper; a postprocessing unit that disconnects a printed sheet of paper in page units, and stores the disconnected sheets of paper in a stack; a deciding unit that determines whether the postprocessing unit has an inversion mechanism that inverts front and back sides of the printed sheets of paper; and a control unit that determines which one of the first and second printing units prints a first print data portion for the first side of the paper and a second print data portion for the second side of the paper in the print data respectively, based on presence or absence of the inversion mechanism. |
Composition for dyeing keratinous fibers containing a paraphenylenediamine substituted by a diazacycloheptane radical |
The subject of the invention is a composition for the oxidation dyeing of keratinous fibres, in particular human hair, comprising an oxidation base of the para-phenylenediamine type, substituted with a diazacycloheptane radical. The invention also relates to the method for dyeing fibres using this composition, and the novel compounds of the type mentioned above. |
1-22 (canceled) 23. A composition for the oxidation dyeing of keratin fibers comprising, in a medium suitable for dyeing, at least one oxidation base chosen from compounds of the following formula (I) and the addition salts thereof: wherein: R1 is chosen from: a halogen atom; and linear and branched C1-C6 hydrocarbon chains, wherein the hydrocarbon chains may be saturated or may comprise at least one bond chosen from double bonds and triple bonds, the hydrocarbon chains may form at least one ring chosen from 3- to 6-membered rings, wherein at least one carbon atom can be replaced by at least one entity chosen from oxygen, nitrogen and sulphur atoms, and an SO2 group, or, when the carbon is terminal, it may be replaced by a halogen atom; provided that the radical R1 does not comprise a peroxide bond, a diazo radical, a nitro radical, or a nitroso radical; n is a number ranging from 0 to 4 inclusive, it being understood that when n is greater than or equal to 2, then the radicals R1 may be identical or different; R2 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated, unsubstituted and substituted with at least one radical chosen from carboxyl radicals, alkylcarbonyl radicals, alkoxycarbonyl radicals, carbamoyl radicals, mono- and dialkylcarbamoyl radicals, saturated and unsaturated, nitrogen- oxygen- and sulphur-comprising heterocyclic radicals comprising 4, 5, 6 or 7, identical or different, atoms; alkyl radicals which may be unsaturated, substituted at least at the 2-position by at least one radical chosen from hydroxyl radicals, alkoxy radicals, amino radicals, mono- and dialkylamino radicals, thiol radicals and halogen atoms; an alkylcarbonyl radical; an alkoxycarbonyl radical; a monoalkylcarbamoyl and dialkylcarbamoyl radical; a carbamoyl radical; and a radical R6R7N—C═NR5— wherein R5, R6, and R7, which may be identical or different, are each chosen from hydrogen, C1-C4 alkyl radicals and hydroxyalkyl radicals; R3 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated; a hydroxyl radical; a hydroxyalkyl radical; an alkoxy radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an amino radical; a monoalkylamino and a dialkylamino radical; an aminoalkyl radical, it being possible for the amine to be mono- or disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; R4 is chosen from: alkyl radicals which may be unsaturated; a hydroxyalkyl radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an aminoalkyl radical, wherein the amine may be mono- or disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; and m is a number ranging from 0 to 4 inclusive, it being understood that when m is greater than or equal to 2, then the radicals R4 may be identical or different. 24. The composition according to claim 23, wherein, in R1, the halogen atom is chosen from chlorine and bromine. 25. The composition according to claim 23, wherein R2 is radical R6R7N—C═NR5—, wherein R5 is a hydrogen atom and R6 and R7, which may be identical or different, are each chosen from a hydrogen atom and a methyl radical. 26. The composition according to claim 23, wherein R1 is chosen from alkyl, hydroxyalkyl, aminoalkyl, alkoxy, and hydroxyalkoxy radicals. 27. The composition according to claim 23, wherein R1 is chosen from methyl, hydroxymethyl, 2-hydroxyethyl, 1,2-dihydroxyethyl, methoxy, and 2-hydroxyethoxy radicals. 28. The composition according to claim 23, wherein n is equal to 0 or 1. 29. The composition according to claim 23, wherein R2 is chosen from hydrogen, alkyl radicals, alkyl radicals substituted with at least one entity chosen from saturated and unsaturated, nitrogen-, sulfur-, and oxygen-comprising heterocyclic radicals comprising 4, 5 or 6, identical or different atoms, an alkoxycarbonyl radical, and alkyl radicals substituted at least at the 2-position with at least one hydroxyl radical. 30. The composition according to claim 23, wherein R2 is chosen from a 2-hydroxyethyl radical, a 3-(1-pyrrolidinyl)propyl radical, a methyl radical, an acetyl radical, and hydrogen. 31. The composition according to claim 23, wherein R3 is chosen from hydrogen, alkyl radicals, alkyl radicals substituted with at least one hydroxyl radical, alkyl radicals substituted with at least one amino radical, and carboxyl radicals. 32. The composition according to claim 23, wherein R3 is chosen from a hydrogen atom, hydroxyl radicals, carboxyl radicals, amino radicals, hydroxymethyl radicals, and aminomethyl radicals. 33. The composition according to claim 23, wherein R4 is chosen from hydrogen, alkyl radicals, alkyl radicals substituted with at least one hydroxyl radical, alkyl radicals substituted with at least one amino radical, and carboxyl radicals. 34. The composition according to claim 23, wherein R4 is a hydrogen atom. 35. The composition according to claim 23, wherein the at least one oxidation base of formula (I) is chosen from: 4-(4-Methyl-[1,4]diazepan-1-yl)phenylamine; 4-[1,4]Diazepan-1-yl-phenylamine; 4-[4-(3-Pyrrolidin-1-yl-propyl)-[1,4]diazepan-1-yl]phenylamine; 2-[4-(4-Aminophenyl)-[1,4]diazepan-1-yl]ethanol; 1-[4-(4-Aminophenyl)-[1,4]diazepan-1-yl]ethanone; 4-(4-Aminophenyl)-[1,4]diazepane-1-carboxamidine; 4-(4-Aminophenyl)-N,N-dimethyl-[1,4]diazepane-1-carboxamidine; 1-(4-Aminophenyl)-4-methyl-[1,4]diazepan-6-ol; 1-(4-Aminophenyl)-4-methyl-[1,4]diazepan-6-ylamine; 1-(4-Aminophenyl)-4-(3-pyrrolidin-1-yl-propyl)-[1,4]diazepan-6-ol; 1-(4-Aminophenyl)-4-(3-pyrrolidin-1-yl-propyl)-[1,4]diazepan-6-ylamine; 1-(4-Aminophenyl)-4-(2-hydroxyethyl)-[1,4]diazepan-6-ol; 2-[6-Amino-4-(4-aminophenyl)-[1,4]diazepan-1-yl]ethanol; 2-[4-(4-Aminophenyl)-6-hydroxymethyl-[1,4]diazepan-1-yl]ethanol; 2-Methyl-4-(4-methyl-[1,4]diazepan-1-yl)phenylamine; 4-[1,4]Diazepan-1-yl-2-methylphenylamine; 2-Methyl-4-[4-(3-pyrrolidin-1-yl-propyl)-[1,4]diazepan-1-yl]phenylamine; 2-[4-(4-Amino-3-methylphenyl)-[1,4]diazepan-1-yl]ethanol; 1-[4-(4-Amino-3-methylphenyl)-[1,4]diazepan-1-yl]ethanone; 4-(4-Amino-3-methylphenyl)-[1,4]diazepane-1-carboxamidine; 4-(4-Amino-3-methylphenyl)-N,N-dimethyl-[1,4]diazepane-1-carboxamidine; 1-(4-Amino-3-methylphenyl)-4-methyl-[1,4]diazepan-6-ol; 1-(4-Amino-3-methylphenyl)-4-methyl-[1,4]diazepan-6-ylamine; 1-(4-Amino-3-methylphenyl)-4-(3-pyrrolidin-1-yl-propyly[1,4]diazepan-6-ol; 1-(4-Amino-3-methylphenyl)-4-(3-pyrrolidin-1-yl-propyly[1,4]diazepan-6-ylamine; 1-(4-Amino-3-methylphenyl)-4-(2-hydroxyethyl)-[1,4]diazepan-6-ol; 2-[6-Amino-4-(4-amino-3-methylphenyl)-[1,4]diazepan-1-yl]ethanol; and 2-[4-(4-Amino-3-methylphenyl)-6-hydroxymethyl-[1,4]diazepan-1-yl]ethanol. 36. The composition according to claim 23, wherein the at least one oxidation base of formula (I) is present in an amount ranging from 0.001% to 10% by weight, relative to the weight of the composition. 37. The composition according to claim 36, wherein the at least one oxidation base of formula (I) is present in an amount ranging from 0.005% to 6% by weight, relative to the total weight of the composition. 38. The composition according to claim 23, further comprising at least one coupler chosen from meta-phenylenediamines, meta-aminophenols, meta-diphenols, naphthalene couplers, heterocyclic couplers and the addition salts thereof. 39. The composition according to claim 23, further comprising at least one additional oxidation base chosen from para-phenylenediamines, bisphenylalkylenediamines, para-aminophenols, ortho-aminophenols, heterocyclic bases, and the addition salts thereof. 40. The composition according to claim 23, further comprising at least one direct dye. 41. A method for the oxidation dyeing of keratin fibers comprising, applying to the fibers at least one dyeing composition comprising, in a medium suitable for dyeing, at least one oxidation base chosen from compounds of the following formula (I) and the addition salts thereof: wherein: R1 is chosen from: a halogen atom; and linear and branched C1-C6 hydrocarbon chains, wherein the hydrocarbon chains may be saturated or may comprise at least one bond chosen from double bonds and triple bonds, the hydrocarbon chains may form at least one ring chosen from 3- to 6-membered rings, wherein at least one carbon atom can be replaced by at least one entity chosen from oxygen, nitrogen and sulphur atoms, and an SO2 group, or, when the carbon is terminal, it may be replaced by a halogen atom; provided that the radical R1 does not comprise a peroxide bond, a diazo radical, a nitro radical, or a nitroso radical; n is a number ranging from 0 to 4 inclusive, it being understood that when n is greater than or equal to 2, then the radicals R1 may be identical or different; R2 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated, unsubstituted and substituted with at least one radical chosen from carboxyl radicals, alkylcarbonyl radicals, alkoxycarbonyl radicals, carbamoyl radicals, mono- and dialkylcarbamoyl radicals, saturated and unsaturated, nitrogen- oxygen- and sulphur-comprising heterocyclic radicals comprising 4, 5, 6 or 7, identical or different, atoms; alkyl radicals which may be unsaturated, substituted at least at the 2-position by at least one radical chosen from hydroxyl radicals, alkoxy radicals, amino radicals, mono- and dialkylamino radicals, thiol radicals and halogen atoms; an alkylcarbonyl radical; an alkoxycarbonyl radical; a monoalkylcarbamoyl and dialkylcarbamoyl radical; a carbamoyl radical; and a radical R6R7N—C═NR5— wherein R5, R6, and R7, which may be identical or different, are each chosen from hydrogen, C1-C4 alkyl radicals and hydroxyalkyl radicals; R3 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated; a hydroxyl radical; a hydroxyalkyl radical; an alkoxy radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an amino radical; a monoalkylamino and a dialkylamino radical; an aminoalkyl radical, it being possible for the amine to be mono- and disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; R4 is chosen from: alkyl radicals which may be unsaturated; a hydroxyalkyl radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an aminoalkyl radical, wherein the amine may be mono- or disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; and m is a number ranging from 0 to 4 inclusive, it being understood that when m is greater than or equal to 2, then the radicals R4 may be identical or different. and developing the color with the aid of at least one oxidizing agent. 42. The method according to claim 41, wherein the at least one oxidizing agent is chosen from hydrogen peroxide, urea peroxide, alkali metal bromates, persalts, peracids and oxidase enzymes. 43. The method according to claim 41, wherein the at least one oxidizing agent is mixed at the time of use with the at least one dyeing composition. 44. The method according to claim 41, wherein the at least one oxidizing agent is applied in the form of at least one oxidizing composition simultaneously to or sequentially with the at least one dyeing composition to the fibers. 45. A multicompartment device or kit comprising, at least one first compartment comprising at least one dyeing composition comprising, in a medium suitable for dyeing, at least one oxidation base chosen from compounds of the following formula (I) and the addition salts thereof: wherein: R1 is chosen from: a halogen atom; and linear and branched C1-C6 hydrocarbon chains, wherein the hydrocarbon chains may be saturated or may comprise at least one bond chosen from double bonds and triple bonds, the hydrocarbon chains may form at least one ring chosen from 3- to 6-membered rings, wherein at least one carbon atom can be replaced by at least one entity chosen from oxygen, nitrogen and sulphur atoms, and an SO2 group, or, when the carbon is terminal, it may be replaced by a halogen atom; provided that the radical R1 does not comprise a peroxide bond, a diazo radical, a nitro radical, or a nitroso radical; n is a number ranging from 0 to 4 inclusive, it being understood that when n is greater than or equal to 2, then the radicals R1 may be identical or different; R2 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated, unsubstituted and substituted with at least one radical chosen from carboxyl radicals, alkylcarbonyl radicals, alkoxycarbonyl radicals, carbamoyl radicals, mono- and dialkylcarbamoyl radicals, saturated and unsaturated, nitrogen- oxygen- and sulphur-comprising heterocyclic radicals comprising 4, 5, 6 or 7, identical or different, atoms; alkyl radicals which may be unsaturated, substituted at least at the 2-position by at least one radical chosen from hydroxyl radicals, alkoxy radicals, amino radicals, mono- and dialkylamino radicals, thiol radicals and halogen atoms; an alkylcarbonyl radical; an alkoxycarbonyl radical; a monoalkylcarbamoyl and dialkylcarbamoyl radical; a carbamoyl radical; and a radical R6R7N—C═NR5— wherein R5, R6, and R7, which may be identical or different, are each chosen from hydrogen, C1-C4 alkyl radicals and hydroxyalkyl radicals; R3 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated; a hydroxyl radical; a hydroxyalkyl radical; an alkoxy radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an amino radical; a monoalkylamino and a dialkylamino radical; an aminoalkyl radical, it being possible for the amine to be mono- and disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; R4 is chosen from: alkyl radicals which may be unsaturated; a hydroxyalkyl radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an aminoalkyl radical, wherein the amine may be mono- or disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; and m is a number ranging from 0 to 4 inclusive, it being understood that when m is greater than or equal to 2, then the radicals R4 may be identical or different and at least one second compartment comprising at least one oxidizing composition. 46. A colored product obtained by oxidative condensation of at least one dyeing composition comprising, in a medium suitable for dyeing, at least one oxidation base chosen from compounds of the following formula (I) and the addition salts thereof: wherein: R1 is chosen from: a halogen atom; and linear and branched C1-C6 hydrocarbon chains, wherein the hydrocarbon chains may be saturated or may comprise at least one bond chosen from double bonds and triple bonds, the hydrocarbon chains may form at least one ring chosen from 3- to 6-membered rings, wherein at least one carbon atom can be replaced by at least one entity chosen from oxygen, nitrogen and sulphur atoms, and an SO2 group, or, when the carbon is terminal, it may be replaced by a halogen atom; provided that the radical R1 does not comprise a peroxide bond, a diazo radical, a nitro radical, or a nitroso radical; n is a number ranging from 0 to 4 inclusive, it being understood that when n is greater than or equal to 2, then the radicals R1 may be identical or different; R2 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated, unsubstituted and substituted with at least one radical chosen from carboxyl radicals, alkylcarbonyl radicals, alkoxycarbonyl radicals, carbamoyl radicals, mono- and dialkylcarbamoyl radicals, saturated and unsaturated, nitrogen- oxygen- and sulphur-comprising heterocyclic radicals comprising 4, 5, 6 or 7, identical or different, atoms; alkyl radicals which may be unsaturated, substituted at least at the 2-position by at least one radical chosen from hydroxyl radicals, alkoxy radicals, amino radicals, mono- and dialkylamino radicals, thiol radicals and halogen atoms; an alkylcarbonyl radical; an alkoxycarbonyl radical; a monoalkylcarbamoyl and dialkylcarbamoyl radical; a carbamoyl radical; and a radical R6R7N—C═NR5— wherein R5, R6, and R7, which may be identical or different, are each chosen from hydrogen, C1-C4 alkyl radicals and hydroxyalkyl radicals; R3 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated; a hydroxyl radical; a hydroxyalkyl radical; an alkoxy radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an amino radical; a monoalkylamino and a dialkylamino radical; an aminoalkyl radical, it being possible for the amine to be mono- and disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; R4 is chosen from: alkyl radicals which may be unsaturated; a hydroxyalkyl radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an aminoalkyl radical, wherein the amine may be mono- or disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; and m is a number ranging from 0 to 4 inclusive, it being understood that when m is greater than or equal to 2, then the radicals R4 may be identical or different. 47. A para-phenylenediamine derivative of formula (I) and the addition salts thereof: wherein: R1 is chosen from: a halogen atom; and linear and branched C1-C6 hydrocarbon chains, wherein the hydrocarbon chains may be saturated or may comprise at least one bond chosen from double bonds and triple bonds, the hydrocarbon chains may form at least one ring chosen from 3- to 6-membered rings, wherein at least one carbon atom can be replaced by at least one entity chosen from oxygen, nitrogen and sulphur atoms, and an SO2 group, or, when the carbon is terminal, it may be replaced by a halogen atom; provided that the radical R1 does not comprise a peroxide bond, a diazo radical, a nitro radical, or a nitroso radical; n is a number ranging from 0 to 4 inclusive, it being understood that when n is greater than or equal to 2, then the radicals R1 may be identical or different; R2 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated, unsubstituted and substituted with at least one radical chosen from carboxyl radicals, alkylcarbonyl radicals, alkoxycarbonyl radicals, carbamoyl radicals, mono- and dialkylcarbamoyl radicals, saturated and unsaturated, nitrogen- oxygen- and sulphur-comprising heterocyclic radicals comprising 4, 5, 6 or 7, identical or different, atoms; alkyl radicals which may be unsaturated, substituted at least at the 2-position by at least one radical chosen from hydroxyl radicals, alkoxy radicals, amino radicals, mono- and dialkylamino radicals, thiol radicals and halogen atoms; an alkylcarbonyl radical; an alkoxycarbonyl radical; a monoalkylcarbamoyl and dialkylcarbamoyl radical; a carbamoyl radical; and a radical R6R7N—C═NR5— wherein R5, R6, and R7, which may be identical or different, are each chosen from hydrogen, C1-C4 alkyl radicals and hydroxyalkyl radicals; R3 is chosen from: a hydrogen atom; alkyl radicals which may be unsaturated; a hydroxyl radical; a hydroxyalkyl radical; an alkoxy radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an amino radical; a monoalkylamino and a dialkylamino radical; an aminoalkyl radical, it being possible for the amine to be mono- and disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; R4 is chosen from: alkyl radicals which may be unsaturated; a hydroxyalkyl radical; an alkoxyalkyl radical; an alkylcarbonyl radical; a hydroxyalkoxyalkyl radical; an aminoalkyl radical, wherein the amine may be mono- or disubstituted with at least one identical or different radical chosen from alkyl, acetyl and hydroxyalkyl radicals; a hydroxy- and aminoalkyl radical; a carboxyl radical; a carboxyalkyl radical; a carbamoyl radical; a carbamoylalkyl radical; an alkoxycarbonyl radical; and a monoalkylaminocarbonyl and a dialkylaminocarbonyl radical; and m is a number ranging from 0 to 4 inclusive, it being understood that when m is greater than or equal to 2, then the radicals R4 may be identical or different, wherein said para-phenylenediamine derivative is not 4-(4-methyl-[1,4]diazepan-1-yl)phenylamine, 4-(4-methyl-1,4-diazacycloheptan-1-yl)aniline, 3-cyano-4-(4-t-butoxycarbonyl-1,4-cyclodiazacycloheptane)aniline, 4-(4-butoxycarbonyl-1,4-diazacycloheptane)aniline. |
Device for connecting an optical fibre |
A device for connecting an optical fibre has a connector associated with at least one optical fibre and can be inserted into a receptacle so as to realize an optical connection along an optical connection axis at an angle of 20° between the at least one optical fibre and at least one connection component housed in the receptacle. The connector has a main body for holding a first portion of an end part of at least one optical fibre and to let a second portion of the end part of the optical fibre project. A cover is slidably associated with the main body. The cover houses the second portion of the end part of optical fibre and leaves at least one bare end of the second portion of the end part completely uncovered. |
1.-23. (Canceled) 24. A device for connecting an optical fibre, comprising: a connector adapted to be associated with at least one optical fibre and to be inserted into a receptacle along a predetermined insertion direction so as to realise an optical connection along an optical connection axis between said at least one optical fibre and least one connection component housed in the receptacle, wherein said optical connection axis is inclined at an angle of less than 20° with respect to said predetermined insertion direction and wherein said connector comprises: a main body adapted to hold a first portion of an end part of at least one optical fibre and to leave a second portion of said end part of optical fibre projecting; and a cover slidably associated with said main body and mobile between a first operational position, wherein said cover houses said second portion of the end part of optical fibre inside of it, and a second operational position, wherein said cover leaves at least one bare end portion of predetermined length of said second portion of the end part of optical fibre completely uncovered, said cover further comprising at least one opening for the passage of said second portion of the end part of optical fibre, said at least one opening being of a size which is much larger than the transversal size of said second portion of the end part of optical fibre, so as not to interfere with said fibre during the movement of the cover between said first and second operational positions. 25. The device according to claim 24, wherein said predetermined length of said at least one bare end portion of optical fibre is greater than or equal to 0.5 mm. 26. The device according to claim 24, wherein said predetermined length of said at least one bare end portion of optical fibre is shorter than 3 mm. 27. The device according to claim 24, further comprising a lid associated with said cover and mobile between a first operational position, wherein said at least one opening is closed by said lid, and a second operational position, wherein said at least one opening is exposed to the outside. 28. The device according to claim 27, wherein when said cover is in its first operational position said lid is in its first operational position and when said cover is in its second operational position said lid is in its second operational position. 29. The device according to claim 24, wherein said main body comprises at least one guide element for said second portion of the end part of optical fibre. 30. The device according to claim 29, wherein said main body comprises: a base body provided with at least one first seat adapted to house said first portion of the end part of said at least one optical fibre and defining said at least one guide element; a first upper body adapted to be placed over and associated with said base body so as to define, at said first seat, a channel for housing said first portion of the end part of said at least one optical fibre; and at least one element for clamping at least one initial part of said first portion of the end part of optical fibre onto said base body. 31. The device according to claim 30, wherein said base body and first upper body are made of moulded plastic material. 32. The device according to claim 30, wherein said base body comprises an upstream portion provided with a first seat for housing a fibre-optic cable including said at least one optical fibre. 33. The device according to claim 32, wherein said main body comprises a second upper body adapted to be associated with said base body at said upstream portion of the base body and provided with a second cable housing seat intended to cooperate with said first cable housing seat when said second upper body is positioned onto said upstream portion of the base body, to hold in position, in a substantially stable manner, said fibre-optic cable. 34. The device according to claim 24, having a size which allows its stable and precise housing in a cutting machine with a sliding blade for cutting optical fibres. 35. The device according to claim 24, wherein said receptacle comprises: a connector housing seat extending along said predetermined insertion direction; a member for guiding said connector in said housing seat; a device for releasably blocking said connector in said housing seat; and a device for aligning said at least one second portion of the end part of optical fibre with said at least one connection component along said optical connection axis. 36. The device according to claim 35, wherein said connector housing seat comprises an abutment surface which is active on said cover when the connector is inserted into the receptacle so as to move said cover from said first operational position to said second operational position. 37. The device according to claim 35, wherein said alignment device comprises a base structure, said at least one connection component being integrally associated therewith, and comprising at least one high precision groove extending parallel to said optical connection axis and adapted to house, in a condition of optical alignment with said at least one connection component along said optical connection axis, said end portion of predetermined length of said second portion of end part of optical fibre. 38. The device according to claim 37, wherein said at least one high precision groove is inclined with respect to said optical connection axis at an angle equal to 1°-3°. 39. The device according to claim 37, further comprising a fibre blocking member integrally associated with said device for releasably blocking the connector and intended to cooperate with said base structure of the alignment device when the connector is inserted into the receptacle to hold, in a substantially stable manner, said end portion of predetermined length of said second portion of the end part of optical fibre in said at least one high precision groove. 40. The device according to claim 39, wherein said fibre blocking member comprises an element made of soft material and adapted to come into contact, when the connector is inserted into the receptacle, with said end portion of predetermined length of said second portion of the end part of optical fibre housed in said at least one high precision groove. 41. The device according to claim 35, wherein said alignment device comprises at least one ferule integrally associated with said connection component and provided with a calibrated hole extending parallel to said optical connection axis and adapted to receive said end portion of predetermined length of said second portion of the end part of optical fibre so that said fibre is substantially stable and in a condition of optical alignment with said connection component along said optical connection axis. 42. A connector for optical fibres, comprising: a main body adapted to hold a first portion of an end part of at least one optical fibre and to let a second portion of said end part of optical fibre project; a cover slidably associated with said main body and mobile between a first operational position, wherein said cover houses said second portion of the end part of optical fibre inside of it, and a second operational position, wherein said cover leaves at least one portion of a length greater than 3 mm of said second portion of the end part of optical fibre completely uncovered, said at least one portion of said second portion of the end part of the optical fibre being provided with a bare end portion, said cover further comprising at least one opening for the passage of said second portion of the end part of optical fibre; and an elastic element interposed between said main body and said cover and such as to keep, in rest state, said cover in said first operational position, said at least one opening being of a size which is much larger than the transversal size of said second portion of the end part of optical fibre, so as not to interfere with said fibre during the movement of the cover between said first and second operational positions. 43. A method for manufacturing a connector for optical fibres, comprising the following steps: providing a main body adapted to hold a first portion of an end part of at least one optical fibre and to let a second portion of said end part of optical fibre project; providing a cover adapted to slide on said main body between a first operational position, wherein said cover houses said second portion of the end part of optical fibre inside of it, and a second operational position, wherein said cover leaves at least one portion of a length greater than 3 mm of said second portion of the end part of optical fibre completely uncovered, said at least one portion of said second portion of the end part of the optical fibre being provided with a bare end portion; providing an elastic element between said main body and said cover, said elastic element being such as to keep, in rest state, said cover in said first operational position; and providing in said cover at least one opening for the passage of fibre, said opening being of a size which is much larger than the size of said second portion of the end part of optical fibre, so as not to interfere with said fibre during the movement of the cover between said first and second operational positions. 44. A method for terminating a fibre-optic cable comprising at least one optical fibre with a central portion made of glass material and an outer coating made of acrylate, said cable further comprising at least one plastic tube housing said at least one optical fibre, a plurality of longitudinal Kevlar™ fibres arranged around said at least one plastic tube and a plastic outer sheath, comprising the steps of: providing at least one part of fibre of predetermined length by successively removing the plastic outer sheath, the longitudinal Kevlar™ fibres and said at least one plastic tube from a free end part of predetermined length of said cable; inserting said at least one part of fibre of predetermined length in at least one fibre housing channel formed in a main body of a connector for optical fibres so as to house a first portion of said part of fibre of predetermined length in said housing channel and to let a second portion of said part of fibre of predetermined length project, said connector for optical fibres further comprising a cover slidably associated with said main body and mobile between a first operational position, wherein said cover houses said second portion of the end part of optical fibre inside of it, and a second operational position, wherein said cover leaves at least one portion of a length greater than 3 mm of said second portion of the end part of optical fibre completely uncovered, said at least one portion of said second portion of the end part of the optical fibre being provided with a bare end portion, said cover comprising at least one opening for the passage of fibre, said opening being of a size which is much larger than the size of said second portion of the end part of optical fibre, so as not to interfere with said fibre during the movement of the cover between said first and second operational positions, said connector for optical fibres further comprising an elastic element operationally interposed between said main body and said cover and such as to keep, in rest state, said cover in said first operational position; positioning a part of cable in an appropriate seat provided in the main body upstream of said fibre housing channel; clamping said part of fibre of predetermined length with respect to said main body; blocking said part of cable with respect to said main body; moving said cover from said first to said second operational position so as to expose said end portion of predetermined length of said second portion of the end part of optical fibre; and cutting said end portion of predetermined length of said second portion of the end part of optical fibre at one of its cutting sections. 45. A fibre-optic communication line, comprising at least one cable including at least one optical fibre, said at least one cable being terminated at at least one of its free ends with a connector for optical fibres, comprising: a main body adapted to hold a first portion of an end part of at least one optical fibre and to let a second portion of said end part of optical fibre project; a cover slidably associated with said main body and mobile between a first operational position, wherein said cover houses said second portion of the end part of optical fibre inside of it, and a second operational position, wherein said cover leaves at least one portion of a length greater than 3 mm of said second portion of the end part of optical fibre completely uncovered, said at least one portion of said second portion of the end part of the optical fibre being provided with a bare end portion, said cover further comprising at least one opening for the passage of said second portion of the end part of optical fibre; and an elastic element interposed between said main body and said cover and such as to keep, in rest state, said cover in said first operational position, said at least one opening being of a size which is much larger than the transversal size of said second portion of the end part of optical fibre, so as not to interfere with said fibre during the movement of the cover between said first and second operational positions. 46. A distribution network comprising at least two distribution lines, each distribution line being a fibre-optic communication line comprising at least one cable including at least one optical fibre, said at least one cable being terminated at at least one of its free ends with a connector for optical fibres, comprising: a main body adapted to hold a first portion of an end part of at least one optical fibre and to let a second portion of said end part of optical fibre project; a cover slidably associated with said main body and mobile between a first operational position, wherein said cover houses said second portion of the end part of optical fibre inside of it, and a second operational position, wherein said cover leaves at least one portion of a length greater than 3 mm of said second portion of the end part of optical fibre completely uncovered, said at least one portion of said second portion of the end part of the optical fibre being provided with a bare end portion, said cover further comprising at least one opening for the passage of said second portion of the end part of optical fibre; and an elastic element interposed between said main body and said cover and such as to keep, in rest state, said cover in said first operational position, said at least one opening being of a size which is much larger than the transversal size of said second portion of the end part of optical fibre, so as not to interfere with said fibre during the movement of the cover between said first and second operational positions; and a branching unit associated with each of said distribution lines. |
Wall prosthesis that can be implanted in the center of a wound to reinforce abdominal wall closure |
The invention relates to a prosthesis that can be implanted in the centre of wall wound scarring. The inventive prosthesis, which is intended for use in abdominal surgery, is provided with a geometric shape in the form of sheets that converge in dihedral angles. The sheets or planes are made from a synthetic biotolerated material in porous form with large pores. One of said planes is inserted into the centre of the scarring between the two aponeurotic surfaces to be joined. The other plane(s) of the prosthesis, which is perpendicular to the aforementioned plane, is arranged so as to overlap the aponeurotic edges of the edge of the section. The proliferation obtained around the prosthesis, in the scarring centre and close thereto, provides stress resistance greater than that obtained in standard closures, thereby greatly reducing the risk of hernias caused by a badly healed wound. |
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