A tissue repair material derived from fish skin and manufacturing method thereof is applied to provide the tissue repair material suitable for use as a patch, a cover, a carrier, a scaffold, an implant or a reagent in various tissues. The tissue repair material has collagens to improve the wounded tissue repair, and has particular characters for desired tissue repair application. Furthermore, so far the factors of the terrestrial animal transmitted disease (caused by virus) do not survive on the tissue repair material derived from fish skin.

Technical Field
The instant disclosure relates to a tissue repair material, and particularly relates to a tissue repair material derived from fish skin and manufacturing method thereof, where the tissue repair material is suitable for use as a patch, a cover, a carrier, a scaffold, an implant or a reagent in various tissues.
Related Art
Tissue engineering that involves the incorporation of a biomaterial with biologics and/or pharmaceutics and upon implantation in a patient will stimulate angiogenesis, tissue integration, and/or tissue remodeling. The biomaterial is a tissue repair material which is synthetic and biocompatible and is used to construct artificial organs, rehabilitation devices, or prostheses and replace natural body tissues.
Collagens are important components in tissue repair material, and have the capabilities of improving tissue growing, improving wound healing, and etc. Additionally, the applications of the tissue repair material with collagens continue to expand. Consequently, it is one of topic researches that the tissue repair material with collagens which has higher medical values and wider diversities is developed for various applications.
Nowadays, the tissue repair materials with collagens are autologous or originate from the donor or the animals. However, in certain diseases, the autologous tissue repair materials cannot be acquired or the acquired tissue repair materials are not enough to use. Furthermore, these tissue repair materials originating from the donor has the difficulty in the supply of donor, the complicated preparing procedure, relatively costly, risk of immunological rejection, and etc. And, these tissue repair materials originating from the animals has the complicated preparing procedure, relatively costly, risk of immunological rejection and terrestrial animal transmitted disease.

Tissue repair material derived from fish skin and manufacturing method thereof


A system and method for use in an implantable cardiac stimulation device permits automatic induction of a tachyarrhythmia of a heart to permit the performance of an electrophysiological test of the heart. A pulse generator repeatedly delivers a group of first and second sets of pacing pulses to a chamber of the heart. The pacing pulses are separated in time by interpulse intervals to overdrive pace a chamber of the heart. A processor, coupled to the pulse generator, varies the second set of interpulse intervals according to a predetermined protocol after each group of pacing pulses is delivered to the chamber of the heart. The successive groups of pacing pulses are delivered to the heart until the tachyarrhythmia is induced.

Frequently, a clinician must perform electrophysiological studies in patients having implanted permanent pacemakers or cardioverters/defibrillators to determine accurately the patient's pathological condition, cardiovascular characteristics and other information needed in order to prescribe a particular therapeutic treatment for the patient. Such studies often require the inducement of a tachyarrhythmia of the heart such as ventricular tachycardia.
Originally, these studies were invasive because they required the surgical insertion of instruments such as temporary intracardiac pacing catheters into the patient. Thus, these studies were accompanied by some risk and preferably were performed in hospitals. Therefore, these studies were not only expensive, but also time consuming and caused patient anxiety.
In order to avoid medical risks to the patient and hospitalization, non-invasive programmed stimulation (NIPS) procedures have been developed to permit the electrophysiological studies to be performed in a physician's office during routine follow-up visits.
These procedures utilize the implanted cardiac stimulation device and an external programmer. By virtue of the presence of a permanent pacing/defibrillation lead associated with the implanted cardiac stimulation device, the need for placement of a temporary intracardiac pacing catheter is eliminated. Typically, NIPS procedures consist generally of the application of premature electrical pulses at precise intervals to the myocardium of the patient's heart by the implanted cardiac stimulation device and its associated lead or leads. The implanted device applies the stimulation pulses under commands from the external programmer which is manually controlled by the clinician. Prior to the procedure, the clinician manually defines the intervals between the successive stimulation pulses on the programmer. After the implanted device applies the succession of stimulation pulses to the heart, the clinician then, on the programmer, manually observes results such as tachyarrhythmia induction or lack of capture by the stimulation pulses. Thereafter, the clinician alters the stimulation pulse intervals and causes the programmer to initiate another application of the successive stimulation pulses to the patient's heart by the implanted cardiac stimulation device. As a result, NIPS studies, as currently constituted, are tedious to administer, consume considerable time, and lead to considerable cost to the electrophysiology laboratory.

Implantable cardiac stimulation device including a system for and method of automatically inducing a tachyarrhythmia


A pull-out wand is disclosed for use with a water delivery device. The pull-out wand may include one or more sensors, such as a touch sensor and/or a proximity sensor.

The present invention relates generally to a pull-out wand for use with a faucet or other water delivery device, and in particular to a pull-out wand having one or more sensors coupled to the pull-out wand.
Pull-out wands are known. Further, proximity and touch sensors are known for use with faucets.
In an exemplary embodiment of the present disclosure, a water delivery device in fluid communication with at least one source of water positioned below a mounting deck is provided. The water delivery device comprising a base portion in fluid communication with the at least one source of water and a pull-out wand portion in fluid communication with the base portion. The pull-out wand portion having at least one water output. The pull-out wand portion being moveably between a first position proximate to the base portion and a second position spaced apart from the base portion. The water delivery device further comprising a sensor coupled to the pull-out wand portion and a valve interposed between the at least one water output of the pull-out wand portion and the at least one source of water. The valve being operable to permit communication of water provided by the at least one source of water to the at least one water output of the pull-out wand portion in a first configuration and to prevent communication of water provided by the at least one source of water to the at least one water output in a second configuration. The water delivery device further comprising a controller operably coupled to the sensor and operably coupled to the valve. The controller causes the valve to be in the first configuration in response to a first indication from the sensor.
In another exemplary embodiment of the present disclosure, a pull-out wand for use with a base portion having an associated controller which controls a flow of fluid through the base portion is provided. The pull-out wand comprising a housing moveable between a first position proximate the base portion and a second position spaced apart from the base portion; a waterway within the housing in fluid communication with the base portion; and a sensor supported by the housing. The sensor operably coupled to the associated controller of the base portion.
In a further exemplary embodiment of the present disclosure, a water delivery device for use by a user is provided. The water delivery device being in fluid communication with at least one source of water positioned below a mounting deck. The water delivery device comprising a base portion in fluid communication with the at least one source of water; a pull-out wand portion in fluid communication with the base portion and having at least one water output, a valve interposed between the at least one water output of the pull-out wand portion and the at least one source of water, an in water sensor adapted to detect if the user is contacting the water exiting the at least one water output of the pull-out wand portion, and a controller operably coupled to the in water sensor and operably coupled to the valve. The pull-out wand portion being moveably between a first position proximate to the base portion and a second position spaced apart from the base portion. The valve being operable to permit communication of water provided by the at least one source of water to the at least one water output of the pull-out wand portion in a first configuration and to prevent communication of water provided by the at least one source of water to the at least one water output in a second configuration. The controller causing the valve to remain in the first configuration in response to the in water sensor detecting the user being in contact with the water exiting the at least one water output of the pull-out wand portion.
Additional features and advantages of the present invention will become apparent to those skilled in the art upon consideration of the following detailed description of the illustrative embodiment exemplifying the best mode of carrying out the invention as presently perceived.

Fluid delivery device with an in-water capacitive sensor


A method is described for making a change in an object oriented relational database in which objects that represent items of persistent data are related to other objects that represent items of persistent data. The method comprises referring to a backward reference contained in a first object that does not have visibility to a second object that the first object is in a unidirectional relationship with. The first object represents a first item of persistent data. The second object represents another item of persistent data. The backward reference identifies the second object. The method also comprises removing the unidirectional relationship, adding a new unidirectional relationship from a third object to the first object, and modifying the backward reference to identify the third object rather than the second object. A cascade delete method that uses a backward reference is also described.

Relational databases are used to define relationships between items of persistent data. For example, FIG. 1a shows a simplistic arrangement of relationships defined within a relational database. Here, an object oriented environment is assumed where first, second, third and fourth objects are used to represent "Customer A", "Customer B", "Order 1" and "Order 2", respectively.
The relational database entries of FIG. 1a can be used, for example, to keep a record of the items purchased for specific customers. Specifically, specific items purchased by Customer A are listed in the "Order 1" object; and, specific items purchased by Customer B are listed in the "Order 2" object. Here, the Customer A object "represents" Customer A and can be assumed to keep various items that identify Customer A (e.g., name, address, phone number, etc.); and, the Customer B object "represents" Customer B and can be assumed to keep various items that identify Customer B (e.g., name, address, phone number, etc.).
Notions of "navigability" come into play in the design of a relational database. Navigability defines the ordered flow in which elements of data within a relational database can be accessed. For example, according to the simplistic relational database entries observed in FIG. 1a, purchased items listed in "Order 1" can be retrieved with the identity of "Customer A" (and purchased items listed in "Order 2" can be retrieved with the identity of "Customer B")—but—the identity of "Customer A" can not be retrieved from the records of "Order 1" (and the identity of "Customer B" can not be retrieved from the records of "Order 2").
Unidirectional relationships 101, 102 enforce the above policy in which information can be obtained in a first direction of object access flow but not in a second. In a typical application, the Customer A object would include information that defines the unidirectional relationship 101 to Order 1 but the Order 1 object would not include any such information (i.e., only the Customer A object has information that corresponds to relationship 101); and, the Customer B object would include information that defines the unidirectional relationship 101 to Order 2 but the Order 2 object would not include any such information (i.e., only the Customer B object has information that corresponds to relationship 102).
In terms used by those in the software arts, it can be said that customer objects have "visibility" to the order objects; but, the order objects not have "visibility" to the customer objects. Restricting visibility in this manner enforces the unidirectional relationship that is to be implemented into the database. Implementing unidirectional relationships within an object oriented persistent data relational database as described just above can be part of the defined software environment.
For example, the current Java Enterprise Java Bean (EJB) 2.0 specification currently implements a unidirectional relationship by instantiating "accessor" methods in an object having visibility to another object in a unidirectional relationship—but not instantiating such accessor methods in the other object that does not have visibility to the object. A first type of accessor method, referred to as a "set" method, is used to create or change a relationship. A second type of accessor method, referred to as a "get" method, give access to another object from the object that executes the accessor method. The EJB 2.0 specification provides as part of its container managed persistence (CMP) for both types of accessor methods and specifies that objects representing items of persistent data are implemented as "entity beans". CMP is a service by which the persistence management of beans within an EJB container are managed by the contained itself.
Implementing a unidirectional relationship where a first object has "visibility" to a second object but the second object does not have "visibility" to the first object can be inefficient, however, when changes are to be made to existing relationships between existing objects. For example, consider if the working environment of the relational database of FIG. 1a is that: 1) there is to be only one customer per order; and, 2) there are high priority customers and low priority customers where high priority customers are given access to items in shortage over low priority customers.
If Customer A is a high priority customer and Customer B is a low priority customer; and if, Order 2 has every item in Order 1 and includes an item in shortage; then, if Customer A suddenly updates his items for purchase to include the item in shortage, Customer A may be satisfied by "taking" Order 2 from Customer B and dropping Order 1. FIG. 1b represents the resulting object relationships.
Implementing the change, however, can be inefficient because of the lack of visibility that Order 2 had to Customer B in the original set of relationships in FIG. 1a. For example, assume the change is implemented by: 1) first searching through all order objects (which could be many more than those observed in FIG. 1a) to find an order that "matches" the needs of Customer A (to which Order 2 will be a "hit"); and then, 2) establishing the new relationship if the customer of an object identified through 1) is lower priority than Customer A.
Because of the lack of visibility that Order 2 has with respect to Customer B, the second step 2) above can only be accomplished by searching through all customer objects (which again can be many more than those observed in FIG. 1a) to find "the object" (Customer B) that is related 102 to Order 2 so as to resolve the customer priority issue. Thus, as searches can be time consuming, the lack of visibility from Order 2 to Customer B causes changes to be inefficiently implemented when an object whose relationship to be changed (in this case Order 2) is identified before its relationship companion (in this case Customer B); and, the first identified object does not have visibility to the companion.

This patent is subject to a terminal disclaimer.

Cascade delete using a backward reference persistent data target object within a unidirectional relational database relationship


A sign having a changeable array constituted by a plurality of pivotally mounted disks which have opposite bright and dark surfaces. The bright surface of each disk is illuminated by an adjacent light emitter when positioned in a direction of viewing. Light from each light emitter is blocked from illuminating disks other than the adjacent disk in the array.

FIELD OF THE INVENTION
This invention relates to a display element which may be used alone as an indicator but will be more commonly used as a pixel in an array of such elements to form a changeable display sign. The invention will be found particularly useful for signs and displays which must be viewed over wide angles such as bus destination signs.


Apparatus and methods to hold and manipulate dental floss for the removal of plaque and food particles from teeth surfaces. The invention comprises: an elongated handle which may house, contain, and dispense a supply of dental floss; a stationary tong which does not generally move with respect to the handle; a pivotal tong which can be pivoted about a pivot point; means for securing the pivotal tong to the handle to selectively adjust the distance between terminal ends of the stationary tong and the pivotal tong; and means for anchoring the dental floss to the terminal ends of the stationary and pivotal tongs, the dental floss being positioned to span the distance and be pulled taut between the terminal ends of the tongs. Means for guiding the length of dental floss along the stationary and pivotal tongs and/or means for cutting the dental floss may also be provided.

Dental research has clearly identified that the primary causative agents of dental caries and periodontal diseases are the build-up of plaque and growth of oral microflora on the coronal and root surfaces of teeth. Plaque build-up is accelerated by the presence of solid food particles and dissolved food constituents which tend to adhere to teeth surfaces. It is also well established that proper oral hygiene, consisting of regularly scouring plaque from the surfaces of teeth, reduces the occurrence of dental caries and periodontal diseases. Oral hygiene is most effective when it is practiced immediately after eating.
The toothbrush is a widely used implement to scour or clean teeth. Even though brushing with a toothbrush is an effective way to clean dental surfaces which oppose the cheeks, lips and tongue, it is less effective to scour or clean plaque from the interproximal surfaces or interstitial areas of teeth. The interproximal surfaces of teeth are those tooth surfaces which face and often contact other tooth surfaces.
Another popular oral hygiene tool is the toothpick. A toothpick is generally a thin sliver of wood or plastic with tapered ends. One end of the toothpick is urged between the teeth to force out larger food particles which may have become entrapped therebetween.
A variety of dental instruments known as scalers and probes are also commonly used by dentists and oral hygienists. A scaler has a probe end with a cutting surface used to remove tartar from teeth. A dental probe has a pointed end which is usually bent from the longitudinal axis of the handle and is used to locate indentations and cavities in teeth. Many configurations are known but such instruments are designed for use by health professionals and are not suited, or intended, for general public use.
Dental floss is the most commonly used implement to scour or clean the interproximal surfaces of teeth. Dental floss is a strong, thin thread. The respective ends of the dental floss are wound about the index finger on each hand of the user. The free length of floss located between the user's fingers is pulled back and forth through the spaces between the teeth. As the thread is pulled, the surfaces of the thread are urged onto the interproximal surfaces of the teeth, thereby scouring and removing food and plaque from those areas of the teeth.
This method of applying dental floss is very wasteful since the terminal ends of the floss are not used and are discarded. In addition, many people have a great deal of difficulty mastering the techniques of correctly using and administering hand-held dental floss. The manipulation of the floss requires a high order of dexterity. The fingers of the user must be inserted into the mouth which can be unsanitary, cumbersome, uncomfortable, and distasteful. The user's fingers become coated with saliva and it is difficult to maintain a solid grip on the floss. The working area within one's mouth is limited which increases the difficulty in using this method of oral hygiene.
Consequently, many different devices have been created to assist in the manipulation and use of dental floss. The following issued patents describe various devices for holding dental floss: Tesberg (U.S. Pat. No. 3,960,159; issued Jun. 1, 1976); Krivit (U.S. Pat. No. 4,026,308; issued May 31, 1977); Johansson et al. (U.S. Pat. No. 4,041,962; issued Aug. 16, 1977); Johnson (U.S. Pat. No. 4,192,330; issued Mar. 11, 1980); Withers (U.S. Pat. No. 4,404,978; issued Sep. 20, 1983); Morin et al. (U.S. Pat. No. 4,556,074; issued Dec. 3, 1985); Lambert (U.S. Pat. No. 4,706,694; issued Nov. 17, 1987); Bianco (U.S. Pat. No. 4,738,271; issued Apr. 19, 1988); Wisegerber (U.S. Pat. No. 4,788,990; issued Dec. 6, 1988); and Kuo (U.S. Pat. No. 4,790,336; issued Dec. 13, 1988).
The inventor believes that the listed patents taken alone or in combination neither anticipate nor render obvious the present invention. These citations do not constitute an admission that such disclosures are relevant or material to the present claims. Rather, these citations relate only to the general field of the disclosure and are cited as constituting the closest art of which the inventor is aware.


A multi-mode interface circuit for coupling a delta sigma modulator (24) to a processor includes a decoder 20 for decoding mode selection inputs to produce a plurality of control signals controlling an oscillator and a plurality of multiplexers. The oscillator is enabled to produce a first clock signal (INTCLK). The first clock signal or an external clock signal (EXTCLK) is multiplexed as a second clock signal (40) to the input of a code generator circuit (23), causing it to generate a clock input signal (MCLK) applied the delta sigma modulator, third and fourth clock signals (41, 42), and a phase-shift-modulated signal (43) in response to both the second clock signal and a 1-bit data signal (MDAT) produced by the delta sigma modulator. One of the third and fourth clock signals and a ground signal is multiplexed to a clock conductor (13). The phase-shift-modulated signal or the 1-bit data signal is multiplexed onto a data output conductor (14).

The invention relates to a method and circuit for providing convenient, multi-mode interfacing between a delta sigma modulator and another device, such as a digital signal processor (DSP) or a digital filter coupled to a DSP.
Ordinarily, only 2 signals are utilized for communication between a delta sigma modulator and a DSP or other processor or a digital filter such as a decimation filter. For example, one of the signals could be an external clock signal generated by the DSP, and the other signal could be a data output signal generated by the delta sigma modulator. The clock signal is required in addition to the data signal because it is necessary to have a clock signal synchronized to the 1-bit serial data signal generated by a delta sigma modulator in order to convert its 1-bit serial data output signal to a more useful multi-bit word of predetermined resolution.
In the closest prior art, the clock signal and data signal are carried by first and second conductors, respectively, which are directly connected between the delta sigma modulator and a DSP (or other processor, such as a microcontroller). This direct connection technique works effectively and economically if the transducer and delta sigma modulator do not need to be electrically isolated from the DSP. However, in many applications the transducer to be measured is electrically xe2x80x9cfloatingxe2x80x9d at a level that may be hundreds or even thousands of volts above or below the ground level of the DSP or other processor. Consequently, for safety reasons and other reasons, it often is necessary for the transducer and the delta sigma modulator to be electrically isolated from the processor (or a digital filter which would be connected in front of the processor to convert the serial 1-bit data output signal produced by the delta sigma modulator to a more useful multi-bit digital word). The electrical isolation often is accomplished by a pair of optical couplers (or other galvanic isolation devices such as isolation transformers, isolation barrier capacitors, or isolating transistor level shift circuits), one optical coupler being used to couple the clock signal of the delta sigma modulator to or from the processor and the other optical coupler being used to couple the 1-bit serial delta sigma output data signal to an input of the processor. However, it is undesirable to be required to use two optical couplers or two other galvanic isolation devices, because optical couplers and other galvanic isolation devices are very expensive.
Thus, there is an unmet need for a convenient, flexible, inexpensive interface between a delta sigma modulator and a processor (such as a DSP or microcontroller) or a digital filter coupled to an input of the processor.
There also is an unmet need for a way of avoiding the need to use two expensive optical couplers or two other galvanic isolation devices to accomplish analog-to-digital conversion of an analog signal produced by a transducer that is referenced to a first ground voltage and providing the digital output as an input to a processor and/or digital filter that is referenced a second ground voltage which may be very different from the first ground voltage.
There also is an unmet need for a circuit and method that provides synchronized digital signals produced by analog-to-digital conversion of a plurality of analog transducer output signals or the like as inputs to a processor.
There also is an unmet need for a circuit and method for providing communication between a processor and a plurality of delta sigma ADCs coupled to perform analog-to-digital conversion of the outputs of a plurality of transducers or the like using a minimum number of conductors coupled between the processor and the delta sigma ADCs.
Accordingly, it is an object of the invention to provide a multi-mode delta sigma modulator circuit including a flexible interface circuit which is capable of a number of selectable operating modes for coupling its clock input signal and its 1-bit serial data output signal to a digital filter or DSP or other processor for any of a variety of user applications.
It is another object of the invention to provide a multi-mode interface circuit which is capable of a number of selectable operating modes for coupling its clock input signal and its 1-bit serial data output signal to a digital filter or DSP or other processor for any of a variety of user applications.
It is another object of the invention to provide a multi-mode interface circuit which is selectively capable of coupling both clock input information and digital output information of a delta sigma modulator through only a single conductor to a digital filter or DSP or other processor.
It is another object of the invention to provide a multi-mode interface circuit which is capable of coupling both clock input information and digital output information of a delta sigma modulator through only a single optical coupler or other galvanic isolation device to a digital filter or a DSP or other processor, wherein the delta sigma modulator has an analog input coupled to an electrically floating transducer or the like.
Briefly described, and in accordance with one embodiment thereof, the invention provides a multi-mode interface circuit (1) for coupling a delta sigma modulator (24) to a processor or a digital filter (37). The interface circuit (1) includes a decoder 20 for decoding a plurality of mode selection inputs to produce a plurality of control signals (30-32,33-1,33-2). An oscillator (21) is enabled by a first control signal (30) from the decoder to produce a first internal clock signal (INTCLK). A first multiplexer (22) selectively switches the first internal clock signal (INTCLK) or an external clock signal (EXTCLK) onto a first conductor (40) to produce a second internal clock signal (40) thereon. A code generator circuit (23) receives the second internal clock signal (40) and a 1-bit data signal (MDAT) produced by the delta sigma modulator (24), and generates a clock input signal (MCLK) applied to a clock input of the delta sigma modulator (24), a third internal clock signal (41), and a fourth internal clock signal (42). The code generator circuit also generates a phase-shift-modulated signal (43) in response to both the second internal clock signal (40) and the 1-bit data signal(MDAT). A second multiplexer (25) selectively switches one of the third (41) and fourth (42) clock signals, and a predetermined signal onto a clock conductor (13) to produce an output clock signal (CLK) thereon. The second multiplexer (25) also selectively switches the external clock signal (EXTCLK) which, if present, is externally applied to the clock conductor (13), from the clock conductor to an input of the first multiplexer (22). A third multiplexer (26) selectively switches the phase-shift-modulated signal (43) or the 1-bit data signal (MDAT) onto a data output conductor (14) to produce a data output signal (DATA) thereon.
In the described embodiment, the code generator circuit produces the clock input signal (MCLK) in response to the second internal clock signal (40). The code generator circuit includes a divider circuit that divides the clock input signal (MCLK) by 2 to produce the third internal clock signal (41) and divides the clock signal (MCLK) by 4 to produce the fourth internal clock signal (42). The code generator circuit also includes a phase-shift-modulator circuit (23B) that phase-shift-modulates the clock input signal (MCLK) according to the 1-bit data signal (MDAT) to produce the phase-shift-modulated signal (43).
The decoder has first (M0) and second (M1) mode selection inputs. The decoder decodes a first state (0,0) of the first and second mode selection inputs to establish a first mode (Mode 0) wherein a first control signal (30) enables the oscillator (21) to produce the first internal clock signal (INTCLK), a second control signal (31) causes the first multiplexer (22) to switch the first internal clock signal (INTCLK) to the first conductor (40) causing the code generator circuit to produce the clock input signal (MCLK) in response to the first internal clock signal (INTCLK), a third control signal (33-1) causes the second multiplexer (25) to switch the third internal clock signal (41) onto the output clock conductor (13) to produce the output clock signal (CLK) equal to the third internal clock signal (41=INTCLK/2), and a fourth control signal (32) causes the third multiplexer (26) to switch the 1-bit data signal (MDAT) onto the data output conductor (14).
The decoder decodes a second state (0,1) of the first and second mode selection inputs to establish a second mode (Mode 1) wherein the first control signal (30) enables the oscillator (21) to produce the first internal clock signal (INTCLK), the second control signal (31) causes the first multiplexer (22) to switch the first internal clock signal (INTCLK) to the first conductor (40) causing the code generator circuit to produce the clock input signal (MCLK) in response to the first internal clock signal (INTCLK), the third control signal (33-1) causes the second multiplexer (25) to switch the fourth internal clock signal (42) onto the output clock conductor (13) to produce the output clock signal (CLK) equal to the fourth internal clock signal (41), and the fourth control signal (32) causes the third multiplexer (26) to switch the 1-bit data signal (MDAT) onto the data output conductor (14).
The decoder decodes a third state (1,0) of the first and second mode selection inputs to establish a third mode (Mode 2) wherein the first control signal (30) enables the oscillator (21) to produce the first internal clock signal (INTCLK), the second control signal (31) causes the first multiplexer (22) to switch the first internal clock signal (INTCLK) to the first conductor (40) causing the code generator circuit to produce the clock input signal (MCLK) in response to the first internal clock signal (INTCLK), a fifth control signal (33-2) causes the second multiplexer (25) to switch the predetermined signal onto the output clock conductor (13) to cause the output clock signal (CLK) to be equal to the predetermined signal, and the fourth control signal (32) causes the third multiplexer (26) to switch the phase-shift-modulated signal (43) onto the data output conductor (14).
The decoder decodes a fourth state (1,1) of the first and second mode selection inputs to establish a fourth mode (Mode 3) wherein the first control signal (30) disables the oscillator (21), wherein the second control signal (31) causes the first multiplexer (22) to switch the external clock signal (EXTCLK) to the first conductor (40) causing the code generator circuit to produce the clock input signal (MCLK) in response to the external clock signal (EXTCLK), the fifth control signal (33-2) causes the second multiplexer (25) to switch the to the output clock conductor (13) to an input of the first multiplexer (22). The fourth control signal (32) causes the third multiplexer (26) to switch the 1-bit data signal (MDAT) onto the data output conductor (14).

Flexible interface circuit and method for delta sigma A/D converters


The new heteropolysaccharide S-84, prepared by fermentation of an unnamed Pseudomonas species, ATCC 31562 has valuable properties as a thickening, suspending and stabilizing agent in aqueous systems. It is especially useful in formulating explosive gels. Its chemical composition is within the following weight ranges of sugars: 8-15% glucuronic acid; 40-50% glucose; 35-45% mannose; 5-15% rhamnose; and 3-9% arabinose; it also contains 1-5% acetyl and 2-7% pyruvate groups.

Compound S-84 may be prepared by fermentation of a suitable nutrient medium with a hitherto undescribed organism, based on extensive taxonomic studies, which is an unnamed Pseudomonas species. An unrestricted permanent deposit of this organism employed in making our heteropolysaccharide was made with the American Type Culture Collection on Sept. 10, 1979 under Accession No. ATCC 31562.
The following considerations make the assignment of a new Pseudomonas species justified and necessary.


A wide-angle indirect ophthalmoscope that enables an operator to view the retina from the posterior pole to the equator as a single image. The ophthalmoscope has an observable field that can include the entire retina. The ophthalmoscope includes optical fibers built into a contact lens in a circular pattern. The fibers are connected to a light source to illuminate the retina, and the contact lens is utilized for observation. The fibers are maintained in a pre-determined fixed orientation in the lens to produce uniform illumination over the entire retina.

The present invention relates in general to instruments which are used to examine the interior of the eye and in particular to ophthalmoscopes, as they are often called.
Because an observation of the patient's retina is a prerequisite of a diagnosis, ophthalmoscopes play an important role in a clinical examination. However, since the time when ophthalmoscopy became part of every clinical examination, ophthalmologists have been anxious to improve the performance of ophthalmoscopes, and in particular they have been anxious to improve the field of view and the observable field that can be obtained with an ophthalmoscope. In connection with the foregoing, the field of view is the area of the fundus covered by the observer's eye in any given direction of observation, i.e., the area of the fundus that is seen as one view without scanning, and the observable field is the area of the fundus limited by a boundary beyond which the ophthalmoscope is unable to reach, i.e., the total area of the fundus that can be seen in several views by scanning. The effort that has been made to increase both of these fields has been continuous and somewhat successful.
Ophthalmoscopes are classified into two types or groups--direct ophthalmoscopes and indirect ophthalmoscopes. Direct ophthalmoscopes enable a field of view not larger than 12.degree. of arc, which corresponds to a relatively small area of the fundus (4 mm. in diameter) and only enable an observable field hardly reaching the equator. Direct ophthalmoscopes, however, enable a 15-power magnification of the field of view. Thus, although the direct ophthalmoscope only enables a field of view of barely 2 to 3 disk diameters, this instrument is employed when high magnification of that area is desired. In this regard, prior art indirect ophthalmoscopes only enable a three-power magnification of the field of view.
Because of the inability of direct ophthalmoscopes to provide a large field of view, indirect ophthalmoscopes are widely in use. One reason for the use of indirect ophthalmoscopes, besides the ability of this instrument to explore the peripheral retina more efficiently, is their larger field in view. In this regard, the indirect ophthalmoscope extends the field of view that can be provided to about 35.degree. of arc, that is 8 to 10 disk diameters, and by moving the instrument to scan the patient's eye the observable field that results includes the ora serrata.
The parameter limiting the field of view in the indirect ophthalmoscope is the numerical aperture of the condensing lens. The recent improvement of the condensing lens by increasing its numerical aperture up to F/1 permits an increase in the field of view to approximately to 45.degree. with a +20D lens. The numerical aperture F/1 is the approximate limit of the optical performance possible. A slight increase (55.degree.) can still be gained with the +30D lens at the expense of loosing 1/3 magnification.
A distinct disadvantage of all known ophthalmoscopes is that they do not permit a single view of the fundus from the posterior pole to the equator. In order to obtain such a view of the fundus to make accurate determinations of the diseased state, an ophthalmoscopist must scan this portion of the fundus and draw a map of what he sees as he views one area of the retina and then the next. A hand-drawn or photo map is the only image of the area of the retina from the posterior pole to the equator that the ophthalmoscopist has to work with.
The ideal ophthalmoscope or the ideal fundus camera should give a picture of the whole retina from the posterior pole to the ora serrata. Such a picture would eliminate the time consuming and inaccurate mapping of the fundus which is tiring for both the patient and the ophthalmologist. In accordance with one embodiment of the present invention, which includes clad fibers, such mapping is significantly reduced because the instrument enables a single view of the fundus from the posterior pole to the equator. In another embodiment of the invention, which includes "selfoc" fibers, a single view of the entire retina is possible.
Thus, the present invention is a wide-angle ophthalmoscope which includes fiber optic light guides for illuminating the retina. The light guides function in conjunction with a lens to provide a wide-angle view of the field which is uniformly illuminated by the fiber optic light guides.
An illuminator which includes fiber optics to illuminate the retina is disclosed in French Pat. No. 1.583.436 by Raymond Andre Dudragne.
The illuminator disclosed in this French patent, however, is not considered suitable because it provides only a very restricted field of observation and can produce reflections of the illuminating light which can blur the image of the retina. In addition, French Pat. No. 1.583.436 suffers from a serious deficiency in that this patent does not disclose the proper location of the ring of illuminating fibers, nor does it disclose the importance of the angle of inclination of the illuminating fibers. Furthermore, the French patent does not disclose the importance of wide-angle fibers.
In connection with the foregoing, an important aspect of the present invention is the location of the illuminating circular bundle of fibers, the inclination of the fibers at the contact point with the cornea and the numerical aperture of the fibers. These points are amplified below.


There is provided a semiconductor device of low power consumption and high reliability having DTMOS' and substrate-bias variable transistors, and portable electronic equipment using the semiconductor device. On a semiconductor substrate (11), trilayer well regions (12, 14, 16; 13, 15, 16) are formed, and DTMOS' (29, 30) and substrate-bias variable transistors (27, 28) are provided in the shallow well regions (16, 17). Large-width device isolation regions (181, 182, 183) are provided at boundaries forming PNP, NPN or NPNP structures, where a small-width device isolation region (18) is provided on condition that well regions on both sides are of an identical conductive type. Thus, a plurality of well regions of individual conductive types where substrate-bias variable transistors (27, 28) of individual conductive types are provided can be made electrically independent of one another, allowing the power consumption to be reduced. Besides, the latch-up phenomenon can be suppressed.

In order to decrease power consumption in CMOS (Complementary Metal Oxide Semiconductor) circuits using MOSFETs (Metal Oxide Semiconductor Field Effect Transistors), it is most effective to lower the power supply voltage. However, merely lowering the power supply voltage would cause the drive current of MOSFETs to lower, resulting in a lower operating speed of the circuit. This phenomenon is known to become noticeable as the power supply voltage becomes a triple or less of the threshold of the transistor. Although this phenomenon can be prevented by lowering the threshold, doing so would give rise to a problem of increases in off-leak current of MOSFETs. Therefore, the lower limit of the threshold is defined within a range over which the above problem does not occur. This in turn defines the limits of power consumption reduction since the lower limit of the threshold corresponds to the lower limit of the power supply voltage.
Conventionally, there has been proposed a dynamic-threshold operation transistor (hereinafter, referred to as DTMOS) which employs a bulk substrate for alleviation of the above-mentioned problem (Japanese Patent Laid-Open Publications HEI 10-22462, Novel Bulk Threshold Voltage MOSFET (B-DTMOS) with Advanced Isolation (SITOS) and Gate to Shallow-Well Contact (SSS-C) Processes for Ultra Low Power Dual Gate CMOS, H. Kotaki et al., IEDM Tech. Dig., p. 459, 1996). The aforementioned DTMOS has a characteristic of a capability of high drive current with low power supply voltage by virtue of its effective threshold lowering in an ON state. The reason why the effective threshold of DTMOS lowers in the ON state is that the gate electrode and the well region are electrically short-circuited.
The principle of operation of the N-type DTMOS is explained below. It is noted that the P-type DTMOS operates similarly with the polarity reversed. In the N-type MOSFET, when the gate electrode voltage is at a low level (in an OFF state), the P-type well region voltage is also at a low level, the effective threshold has no differences from normal MOSFETs. Therefore, the off current value (off-leak) is the same as in the case of normal MOSFETs.
On the other hand, when the gate electrode voltage is at a high level (in an ON state), the P-type well region voltage is also at a high level, the effective threshold lowers by the substrate-bias effect, so that the drive current increases compared with those of normal MOSFETs. Therefore, large drive current can be obtained with low power supply voltage while low leak current is maintained.
In a DTMOS, the gate electrode and the well region are electrically short-circuited. Therefore, as the gate electrode voltage changes, the well voltage also changes similarly. This accordingly requires the well region of each DTMOS to be mutually electrically isolated from the well regions of its neighboring MOSFETs. For this reason, the well region is made up of a shallow well region and a deep well region which are different in polarity from each other. Furthermore, the shallow well regions of the respective DTMOS' are electrically isolated from one another by a device isolation region.
As a conventional method for suppressing off-leaks in low voltage drive and still obtaining high drive current, there has been a method in which the well-bias voltage is changed between standby and active states (Japanese Patent Laid-Open Publications HEI 6-216346 and 10-340998).
Hereinafter, a MOSFET in which in which the well bias is changed between standby and active states will be expressed as substrate-bias variable transistor.
The principle of operation of the N-type substrate-bias variable transistor is explained below. It is noted that the P-type substrate-bias variable transistor operates similarly with the polarity reversed. In an N-type substrate-bias variable transistor, when the circuit is in an active state, a 0 V or positive voltage (with a source voltage referenced) is applied from a bias generation circuit to the P-type well region. With a positive voltage applied to the P-type well region, the effective threshold lowers due to a substrate-bias effect, and the drive current increases as compared with the case of normal MOSFETs. When the circuit is in a standby state, on the other hand, a negative voltage is applied from the bias generation circuit to the P-type well region. As a result of this, the effective threshold increases due to the substrate-bias effect, and the off-leak decreases as compared with normal MOSFETs or DTMOS'.
Generally, in a circuit using substrate-bias variable transistors, whether the active state or the standby state is effectuated is selected every circuit block. This is because providing the bias generating circuit for each device causes the number of devices and the circuit area to increase considerably. From these reasons, the P-type well region of an N-type MOSFET is common within a circuit block (the case is the same with the N-type well region of a P-type MOSFET). Accordingly, in a circuit block which is in an active state, a 0 V or positive voltage is applied to the well regions of all the N-type MOSFETs, so that the off-leak increases as compared with normal MOSFETs or DTMOS' (the case is the same also with the P-type MOSFETs).
In the circuit using substrate-bias variable transistors, the MOSFETs have to share a well region. For this purpose, the depth of the bottom face of the device isolation region is set deeper than the depth of the junction between the source regions and drain regions of the MOSFETs and their shallow well region and, at the same time, shallower than the depth of the lower end of the well region.
There has been disclosed a technique in which the DTMOS and the substrate-bias variable transistor are combined together to make the best of their respective advantages (Japanese Patent Laid-Open Publication HEI 10-340998).
FIG. 11 shows a cross-sectional view of a device fabricated by this technique. Referring to FIG. 11, there are shown, with reference numerals having the following denotations, a semiconductor P-type substrate 311, an N-type deep well region 312, a P-type deep well region 313, an N-type shallow well region 314, a P-type shallow well region 315, a device isolation region 316, an N-type MOSFET source region 317, an N-type MOSFET drain region 318, a P-type MOSFET source region 319, a P-type MOSFET drain region 320, an N+ diffusion layer 321 for providing contact with an N-type shallow well region, a P+ diffusion layer 322 for providing contact with a P-type shallow well region, a gate insulator 323, a gate electrode 324, a P-type substrate-bias variable transistor 325, an N-type substrate-bias variable transistor 326, an N-type DTMOS 327, a P-type DTMOS 328, a well-bias input 329 for the P-type substrate-bias variable transistor, a well-bias input 330 for the N-type substrate-bias variable transistor, and a fixed bias input 331 for the P-type deep well region. In addition, although not shown, the gate electrode 324 and the P-type shallow well region 315 are electrically short-circuited in the N-type DTMOS 327, and the gate electrode 324 and the N-type shallow well region 314 are electrically short-circuited in the P-type DTMOS 328.
In the DTMOS' 327 and 328, the voltages of the shallow well regions 314 and 315 change with the voltage of the gate electrode 324. In order to prevent changes of the voltages of the shallow well regions 314 and 315 from affecting shallow well regions 314 and 315 of other devices, deep well regions 313 and 312 opposite in conductive type to the shallow well regions 314 and 315 are formed under those shallow well regions 314 and 315. Moreover, a device isolation region 316 is formed at enough depth to electrically isolate shallow well regions 314 and 315 of mutually neighboring devices. By doing so, the shallow well regions 314 and 315 are electrically isolated from shallow well regions 314 and 315 of neighboring devices. Meanwhile, shallow well regions of the substrate-bias variable transistors 326 contained in one circuit block have to be provided in common. Therefore, under the P-type shallow well regions 315 of the N-type substrate-bias variable transistors 326 in FIG. 11, is formed the P-type deep well region 313, which is integrated with a P-type shallow well region 315 to form a common well region. To this P-type common well region 313, 315, a voltage that differs between active and standby states is given via the well-bias input 330 for the N-type substrate-bias variable transistor 326. In order to prevent any effects on devices of other circuit blocks or DTMOS portion, the N-type deep well region 312 is formed further deeper in the substrate, by which the P-type deep well region 313 is electrically isolated. Under the shallow well region 314 of the P-type substrate-bias variable transistor 325 in FIG. 11, is formed the N-type deep well region 312, which is integrated with the N-type shallow well region 314 to form a common well region 312, 314. To this N-type common well region 312, 314, a voltage that differs between active and standby states is given via the well-bias input 329 for the P-type substrate-bias variable transistor 325.
FIGS. 12 and 13 show the procedure of forming the deep well regions 312, 313 of this conventional semiconductor device. As shown in FIG. 12, with photoresist 332 used as a mask, dopant injection for forming the P-type deep well region 313 is performed, and then dopant injection for forming the N-type deep well region 312 further deeper is performed. Next, as shown in FIG. 13, with the photoresist 332 used as a mask, dopant injection for forming the N-type deep well region 312′ is performed. In this case, the depth of the N-type deep well region 312′ is set to a level similar to the depth of the P-type deep well region 313. By these steps, the N-type deep well regions 312 and 312′ are integrated together, by which the P-type deep well region 312 is electrically isolated.
In this way, the substrate-bias variable transistors 325, 326 and the DTMOS' 327, 328 are formed on the same substrate 311, making it possible to realize a circuit making the best of their respective advantages.
As shown in FIG. 11, in the conventional semiconductor device in which the DTMOS' 327, 328 and the substrate-bias variable transistors 325, 326 are combined together (Japanese Patent Laid-Open Publication HEI 10-340998), although the P-type deep well regions 313, 313, can be electrically isolated, the N-type deep well region 312 is common within one substrate 311. Therefore, although the circuit block of the N-type substrate-bias variable transistors 326, 326, . . . can be formed plurally within the same substrate 311, the circuit block of the P-type substrate-bias variable transistor 325, . . . cannot be formed plurally. For this reason, the circuit block of the P-type substrate-bias variable transistors 325, cannot be divided properly into active circuit blocks and standby circuit blocks. For example, when only part of the P-type substrate-bias variable transistors 325, 325, . . . need to be put into active state, the entirety of the P-type substrate-bias variable transistors 325, 325, . . . would come into an active state, causing the leak current to increase. As a result, the power consumption would increase.
Also, the conventional semiconductor device, in which the N-type deep well regions 312 are integrated together within the substrate 311, has a PN junction of a large area comparable to the area of the whole substrate 311. That is, a very large electrostatic capacity is parasitically present. Therefore, changing over between active and standby states in a circuit block of the P-type substrate-bias variable transistors 325, 325, . . . causes the bias of the whole N-type deep well region 312 to change, which in turn causes large amounts of charges to be charged and discharged. As a result, the power consumption increases.
Furthermore, in the foregoing semiconductor device, putting the P-type substrate-bias variable transistors 325, 325, . . . into the active state (i.e., giving a voltage lower than the power supply voltage to the N-type deep well region 312) makes it more likely that a latch-up phenomenon is induced. In an NPNP type structure having routes passing through the N-type shallow well region 314 of the P-type DTMOS 328, the P-type deep well region 313, the N-type deep well region 312 and the P-type shallow well region 315 of the N-type DTMOS 327, a case (undershoot) is discussed where a bias lower than ground voltage is applied to the N-type shallow well region 314 of the P-type DTMOS 328. In the DTMOS 328, in which the gate electrode 324 and the shallow well region 314 are electrically connected to each other, it can occur that a bias not more than the ground voltage is applied to the N-type shallow well region 314 of the P-type DTMOS 328 through the gate electrode 324. In this case, a forward voltage is applied to the junction between the N-type shallow well region 314 of the P-type DTMOS 328 and the P-type deep well region 313, so that electrons are injected into the P-type deep well region 313. The electrons injected into the P-type deep well region 313 reach the N-type deep well region 312, causing the voltage of the N-type deep well region 312 to decrease. With the voltage of the N-type deep well region 312 decreased, holes are injected from the P-type shallow well region 315 of the N-type DTMOS 327 into the N-type deep well region 312. The holes injected into the N-type deep well region 312 reach the P-type deep well region 313, causing the voltage of the P-type deep well region 313 to increase. With the voltage of the P-type deep well region 313 increased, electrons injected from the N-type shallow well region 314 of the P-type DTMOS 328 into the P-type deep well region 313 increase more and more. Through iterations of these steps (with a positive feedback applied), an abnormal current flow in the NPNP structure, giving rise to a latch-up phenomenon. In this connection, if a voltage lower than the power supply voltage has been applied to the N-type deep well region 312 from the beginning (i.e., if the P-type substrate-bias variable transistor 325 has been in an active state), the latch-up phenomenon is more likely to occur. Further, also when the P-type substrate-bias variable transistor 325 comes into a standby state (i.e., also when a voltage higher than the power supply voltage is applied to the N-type deep well region 312), it become more likely that a latch-up phenomenon is induced. In this case, a large inverse-bias is applied to the junction between the P-type shallow well region 315 of the N-type DTMOS 327 and the N-type deep well region 312, as well as to the junction between the P-type deep well region 313 and the N-type deep well region 312. This causes a punch-through to occur between the P-type shallow well region 315 of the N-type DTMOS 327 and the P-type deep well region 313, triggering the occurrence of a latch-up phenomenon in the NPNP structure. As to the route of the latch-up, in addition to the above-mentioned one, such an NPNP structure may also be mentioned as one including a route which passes the drain region 318 of the N-type DTMOS 327, the P-type shallow well region 315 of the N-type DTMOS 327, the N-type deep well region 312 and the P-type deep well region 313. Thus, largely changing bias of the N-type deep well region 312 makes it difficult to control the latch-up phenomenon. As a result, the device reliability deteriorates.

Semiconductor device and portable electronic apparatus


The present invention describes granulocyte-binding antibody constructs, which were derived from the monoclonal mouse antibody MAk BW 250/183 (EP-A1-0 388 914), as well as a process for their preparation and their use.

The present invention describes granulocyte-binding antibody constructs, which were derived from the monoclonal mouse antibody Mab BW 250/183 (EP-A1-0 388 914), as well as a process for their preparation and their use.
The monoclonal antibody Mab BW 250/183 is a monoclonal mouse antibody of the IgG1 kappa type which is directed against the "nonspecific crossreacting antigen"(NCA)95 and against the carcinoembryonic antigen (CEA) and does not have any unwanted cross reactions with normal human tissues apart from the colon mucosa. Its binding to granulocytes and granulocyte precursors in the bone marrow also makes a therapeutic application possible.
Usually, antibodies which were isolated from rodents, in particular mice, are used in the therapeutic application of monoclonal antibodies (Mabs) in humans. However, the repeated administration of high doses of antibodies of non-human (xenogenic) origin can lead to an immune reaction in the patients. After about 10-14 days they develop anti-mouse immunoglobulin antibodies (HAMAs). A therapy of this nature must therefore be discontinued after 10 days and cannot be reinitiated.
In the meantime, success has been achieved, with the aid of molecular biological methods, in altering the xenogeneic antibodies in such a way that they are either no longer or only very weakly immunogenic for humans but nevertheless preserve their antigen-binding properties. To do this, the constant exons (CH.sub.1, H, CH.sub.2, CH.sub.3, CL) in the genes encoding the H and L chains of the xenogeneic Mabs were exchanged, in a first step, for exons from human immunoglobin genes (Boulianne, G. L., Nobumichi Hozumi, and Marc J. Shulman (1984) Nature 321, 643-646). In this way, so-called chimeric antibodies are obtained which are composed of variable domains of xenogeneic origin and human constant regions. These chimeric antibodies still contain about 30% of xenogeneic protein sequences so that when they are used in immunotherapy an immune reaction, even if weaker, is still to be expected in the patients (Bruggemann, M., Greg Winter, Herman Waldmann, and Michael S. Neuberger (1989) J. Exp. Med. 170, 2153-2157).
In order to reduce the immunogenicity of xenogeneic antibodies still further, a technique was developed by G. Winter and M. Neuberger (1986) in which only the CDR loops of the V.sub.L and V.sub.H domains of the xenogenic antibodies are transferred to V.sub.L and V.sub.H domains of human antibodies (Jones, P. T., Paul H. Dear, Jefferson Foote, Michael S. Neuberger, and Greg Winter (1986) Nature 321,522-525), a process which is termed "humanization" and takes place at the level of the V.sub.H and V.sub.L genes. The structural variability of the V.sub.H and V.sub.L domains is in general confined to the CDR loops ["CDR" stands for complementarity-determining regions].
In the present patent application, antibody chains are now described which are specific for human granulocytes and in which at least the CDR regions of the heavy and light chains derive from the mouse Mab BW 250/183 (EP-A1-0 388 914). The remaining parts of molecular constructs prepared thereby, e.g. monoclonal antibodies, are preferably derived from human antibodies.

Granulocyte-binding antibody constructs, their preparation and use


The present invention provides a method of extracting and isolating nucleic acids from a material containing nucleic acids using a nucleic acid-binding particulate carrier. More specifically, the present invention provides a nucleic acid extraction method using a particulate carrier having a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g. According to the method of the invention, nucleic acids can be efficiently extracted from a biological material, in particular a material containing a large amount of contaminants, such as a clinical sample.

The present invention relates to a method of extracting and isolating nucleic acids from a material containing nucleic acids, using a nucleic acid-binding particulate carrier. More specifically, the present invention relates to a nucleic acid extraction method using a porous particulate carrier having a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g.
Recent advances in the genetic engineering and molecular biology fields enable analyses of infections and genetic diseases at the DNA/RNA level. In particular, detection of a trace amount of nucleic acid, which has so far been extremely difficult, becomes readily achievable, and therefore gene analyses are remarkably facilitated, owing to the invention of nucleic acid amplification methods, such as polymerase chain reaction (PCR; Science 230:1350-1354, 1985) and nucleic acid sequence based amplification method (NASBA; Nature 350: 91-92, 1991 and Japanese Patents Nos. 2648802 and 2650159).
However, for detecting a nucleic acid in a biological sample, or amplifying a nucleic acid before detection as required, it is necessary to selectively withdraw nucleic acids from the sample, since biological samples usually contain large amount of constituents other than nucleic acids, such as proteins, lipids and saccharides, which are likely to adversely affect the amplification or detection. Accordingly, manipulations are required to remove contaminants from biological samples and to extract and isolate nucleic acids.
Various techniques have been employed for nucleic acid isolation. Typical examples include techniques in liquid phases, such as the phenol-chloroform extraction method (Biochimica et Biophysica Acta 72:619-629, 1963) and the alkaline SDS method (Nucleic Acid Research 7:1513-1523, 1979). These techniques are widely used on a laboratory scale, but require technological skills and are difficult to carry out with good reproducibility since they use organic solvents that are toxic and difficult to dispose of, such as phenol and chloroform, and use hazardous materials such as sodium hydroxide.
Nucleic acid isolation techniques utilizing nucleic acid-binding carriers include one utilizing glass particles and a sodium iodide solution (Proc. Natl. Acad. Sci. USA 76-2:615-619, 1979), and one utilizing hydroxyapatite (Japanese Unexamined Patent Publication No. 263093/1988). Although these techniques do not employ toxic organic solvents or other toxic substances, but have the problem that they are not suitable for processing many samples at a time and thus require a prolonged period of time, since they involve a number of centrifugation operations.
In short, conventional nucleic acid isolation techniques as mentioned above have the drawback that they use hazardous reagents such as organic solvents and alkalis, or involve centrifugation operations and are unsuitable for processing many samples at a time. Further, the techniques encounter large problems when the extraction and isolation steps are automatized, although automatization of the extraction and isolation of nucleic acids is indispensable for processing a number of samples with good reproducibility and for reducing labor cost.
For automatization of extraction and isolation of nucleic acids, a technique utilizing silica particles and chaotropic ions has been proposed (J. Clinical Microbiology 28-3:495-503, 1990, and Japanese Unexamined Patent Publication No. 289596/1990). The technique comprises mixing a sample with nucleic acid-binding silica particles and chaotropic ions capable of releasing nucleic acids contained in the sample to thereby bind the nucleic acids to silica particles, removing contaminants by washing, and collecting the nucleic acids bound to the silica particles. This technique is advantageous in that it is suitable not only for extraction of DNA but also for extraction of less stable RNA, and capable of giving high-purity nucleic acids. However, the step of washing the particles having bound nucleic acids involves centrifugation, filtration using a filter, etc., and therefore is liable to become complicated when automatized.
For facilitating mixing and washing of the silica particles, Japanese Unexamined Patent Publication No. 19292/1997 discloses a technique comprising magnetizing the silica particles and mixing a sample with the particles utilizing the magnetic field, followed by stirring. In this technique, nucleic acids are bound to the magnetic silica particles, the particles are magnetically separated from the liquid phase, and after washing the particles, the nucleic acids are collected. The steps of the technique become simpler and easier when automatized. However, the influence of contaminants in the sample cannot be excluded by simply employing magnetized particles, and as the result, this technique often shows reduced collection efficiency.
Namely, when a sample containing a large amount of contaminants is used, contaminants in the sample cover the particle surfaces, whereby nucleic acids adsorb to the particle surfaces at a reduced adsorption rate.
To overcome the problem, Japanese Unexamined Patent Publication No. 262387/1999 proposes a technique utilizing a particulate carrier with a larger particle surface area, and Japanese Unexamined Patent Publication No. 178571/1999 proposes a technique wherein a particulate carrier is suspended in a surfactant beforehand in order to preclude adsorption of contaminants. However, it is still difficult to extract a sufficient amount of nucleic acids from samples containing a large amount of contaminants, such as whole blood samples.
The object of the present invention is to solve the above problems by providing a method for efficiently extracting and isolating nucleic acids from a material containing nucleic acids, in particular from a clinical sample or like sample containing a large amount of contaminants.
The present inventors conducted extensive research to achieve the above object, and found that a porous particulate carrier having a pore diameter of at least 50 nm and a pore volume of at least 200 mm3/g is useful for efficiently extracting nucleic acids from a biological material, in particular from a clinical sample or like sample containing a large amount of impurities. The present invention has been accomplished based on this novel finding.
Porous particulate carriers have a pore diameter (i.e., diameter of the openings of the pores) and a pore volume (i.e., interior volume of the pores). The present invention has been accomplished based on the finding that the pore diameter and the pore volume affect the collection efficiency of nucleic acids from a sample.
The present invention provides a method of extracting nucleic acids from a material containing nucleic acids using a nucleic acid-binding particulate carrier, wherein the particulate carrier has a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g.
Specifically, the present invention provides a method of extracting nucleic acids from a material containing nucleic acids, the method comprising the following steps (a) to (c):
(a) mixing the material containing nucleic acids, a nucleic acid-binding particulate carrier having a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g, and a nucleic acid extraction solution for allowing the nucleic acids to adsorb to the particulate carrier, to thereby bind the nucleic acids to the particulate carrier;
(b) separating a composite of the nucleic acids and the particulate carrier from the mixture obtained in Step (a) to remove contaminants; and
(c) eluting and collecting the nucleic acids from the composite of the nucleic acids and the particulate carrier.
The present invention also provides a nucleic acid extraction kit comprising a nucleic acid-binding particulate carrier having a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g, a nucleic acid extraction solution for allowing the nucleic acids to adsorb to the carrier, and a nucleic acid eluate for eluting the nucleic acids from a composite of the nucleic acids and the particulate carrier.
The present invention further provides a nucleic acid detection method comprising extracting nucleic acids by the above method, amplifying a target nucleic acid by amplification reaction and detecting the target nucleic acid.
The following are definitions of terms used herein and descriptions of measurement methods employed herein.
Nucleic Acid-binding Particulate Carrier
The term xe2x80x9cnucleic acid-binding particulate carrierxe2x80x9d means a particulate carrier capable of binding or adsorbing nucleic acids to the particle surfaces.
The particulate carrier for use in the invention is porous, and xe2x80x9cporesxe2x80x9d of the particulate carrier are minute cavities on the particle surfaces, and xe2x80x9cpore volumexe2x80x9d means the interior volume of the pores. xe2x80x9cPore diameterxe2x80x9d means the diameter of the openings of the pores present on the carrier, and, as used herein, is intended to mean the average diameter of the pores, since a pore diameter of a fine powder usually indicates the average pore diameter. xe2x80x9cParticle diameterxe2x80x9d indicates the diameter of particles, assuming that the particles are spherical.
In the present invention, the pore diameter and pore volume of the particulate carrier are analyzed by mercury penetration. Mercury penetration is an analysis based on the physical principle that mercury wets substantially no porous substances on their pore walls, and thus does not penetrate into the pores unless being pressurized. In this analysis, mercury surrounding the analyte is uniformly pressurized from the periphery. As the pressure gradually increases, mercury penetrates first into pores with a relatively large diameter and then gradually into pores with a smaller diameter. Assuming that there is a pore having a cylindrical shape and a radius of r, the force (xcfx80r2P) of penetration of mercury into the pore when applying a pressure P and the force (xe2x88x922xcfx80rxc3x97xcex3 cos xcex8) of repulsion towards mercury are in equilibrium. Thus, mercury penetrates into pores having a radius greater than the radius r calculated according to the following equation:
Pr=xe2x88x922xcex3 cos xcex8xe2x80x83xe2x80x83(1)
wherein P is the pressure, r is the pore radius, xcex3 is the surface tension of mercury, xcex8 is the contact angle of mercury and the analyte (90xc2x0 C. less than xcex8180xc2x0 C.). The above equation (1) is called Washburn equation, and the value of the right side thereof is a constant which is characteristic of the analyte and is equal to the penetration amount of mercury relative to the pressure or the pore radius. The penetration amount of mercury indicates the cumulative volume of pores having a radius greater than the radius r.
The particle diameter is measured by a particle size distribution measuring method utilizing the difference in the rates of precipitation of particles with different diameters in a solution.
It is essential in the present invention that the particulate carrier has a pore diameter of at least 50 nm and a pore volume of at least 200 mm3/g. Use of a particulate carrier having a larger pore diameter and a larger pore volume makes it possible to collect an increased amount of nucleic acids. If the particulate carrier has a pore diameter less than 50 nm or a pore volume less than 200 mm3/g, there arises the problem that nucleic acids are not bound or adsorbed to the particle surfaces, owing to the influence of contaminants in the sample, or other factors. However, particulate carriers having a pore diameter greater than 500 nm and/or a pore volume greater than 5000 mm3/g are not preferable, since a large amount of constituents other than nucleic acids is presumably liable to be collected in the particles, owing to the excessive spaces in the pores. Further, particles having a pore diameter greater than 500 nm are likely to be brittle and fail to maintain their structure, since the pore diameter occupies a considerable part of the pore diameter. The pore diameter is preferably 50 to 500 nm, more preferably 50 to 300 nm particularly preferably 80 to 250 nm. The pore volume is preferably 200 to 5000 mm3/g, more preferably 300 to 3000 mm3/g.
If having a particle diameter less than 0.5 xcexcm, the particulate carrier has excessively high dispersibility, and is difficult to collect for washing or the like. If the particle diameter is larger than 15.0 xcexcm, the particulate carrier has a high density and therefore rapidly precipitates from the mixture, hence undesirable. The particle diameter is preferably 0.5 to 15.0 xcexcm, more preferably 0.5 to 10.0 xcexcm, particularly preferably 1.0 to 5.0 xcexcm.
For example, during formation of a composite of nucleic acids and the magnetic silica particulate carrier, contaminants derived from organisms tend to adhere to the particle surfaces of the magnetic silica particulate carrier and prevent binding of nucleic acids, when blood or like biological sample containing a large amount of contaminants is used. That is, the greater the amount of contaminants, the lower the nucleic acid collection efficiency becomes. For solving this problem, the method of the invention employs a specific particulate carrier which is large in pore diameter and pore volume, so that even if contaminants in the sample adhere to the carrier, they are less likely to affect the binding of nucleic acids. Therefore, the method of the invention achieves a high collection efficiency.
Accordingly, it is necessary in the method of the present invention to use a particulate carrier having a pore diameter of 50 to 500 nm, a pore volume of 200 to 5000 mm3/g and a particle diameter of 0.5 to 15.0 xcexcm.
The term xe2x80x9csurface areaxe2x80x9d as used herein means the total of the surface areas of the particles of the carrier. It is a common practice to express the surface area of a fine powder in a surface area per unit weight (e.g., 1 g), not in a surface area per particle. The surface area per unit weight is called specific surface area. Further, xe2x80x9couter surface areaxe2x80x9d means the area of the outer surface of the particles of the carrier and is expressed as an outer surface area per unit weight (e.g., 1 g), like the surface area.
In the present invention, the surface area and specific surface area are measured by mercury penetration and gas absorption. The outer surface area of the particulate carrier is found by subtracting the surface area inside the particles from the surface area. The surface area inside the particles is calculated from the pore volume.
Consideration is then given to the specific surface area S of pores. Assuming that there are a number n of cylindrical pores having a radius of r and a depth of l, the increase in specific surface area is:
dS=2xcfx80rlxc3x97nxe2x80x83xe2x80x83(2)
and the increase in pore volume is:
dV=xcfx80r2lxc3x97nxe2x80x83xe2x80x83(3).
From the equations (2) and (3),
dS=(2/r)dV.
Thus, the specific surface area S=(2/r)∫dV.
The outer surface area of the particulate carrier is usually at least 5 m2/g, preferably 5 to 500 m2/g, more preferably 20 to 400 m2/g.
The specific surface area of the particulate carrier is usually 5 to 800 m2/g, preferably 10 to 600 m2/g, more preferably 15 to 500 m2/g.
It is preferred that the particulate carrier for use in the present invention comprises silica or its derivative, and it is more preferred that the particulate carrier be magnetic carrier. As used herein, the term xe2x80x9csilicaxe2x80x9d sometimes means xe2x80x9csilica or its derivativexe2x80x9d.
As used herein, the term xe2x80x9csilica or its derivativexe2x80x9d includes SiO2 crystals and silicon oxides of other forms, skeletons of diatomaceous earth composed of SiO2, and amorphous silicon oxides.
xe2x80x9cMagneticxe2x80x9d indicates capability of being temporarily magnetized by a permanent magnet and attracted to the magnet. Magnetic particulate carriers can be easily separated from a solution phase.
It is more desirable that the particulate carrier for use in the present invention be magnetic particulate carrier comprising silica or its derivative (hereinafter referred to as xe2x80x9cmagnetic silica particulate carrierxe2x80x9d). It is particularly desirable that the particles of magnetic silica particulate carrier be composites of fine particles each comprising a superparamagnetic metal oxide covered with silica, the composites having porous surfaces. The particles of the magnetic silica particulate carrier are substantially complete spheres. Nucleic acids are bound to the silica particulate carrier via hydrogen bonds formed between hydroxyl groups on the particle surfaces of the carrier and bases of the nucleic acids.
Superparamagnetic metal oxides are metal oxides that are responsive to a magnetic field variation but are not permanently magnetized, and have a small residual magnetization. The particle size of the superparamagnetic metal oxide for use in the invention is about 10 to 200 nm, preferably about 20 to 100 nm. Preferred superparamagnetic metal oxides include iron oxides such as triiron tetraoxide (Fe3O4) and xcex3-type iron sesquioxide (xcex3-Fe2O3) obtainable by gradually oxidizing triiron tetraoxide. Triiron tetraoxide has small residual magnetization and a preferred surface structure (substantially complete spheres), so that it can be used repeatedly by magnetic separation and redispersion. Magnetic silica particulate carriers containing triiron tetraoxide are stable in weakly acidic aqueous solutions and can be stored over a period of 2 years or more.
The magnetic silica particulate carrier for use in the invention contains the superparamagnetic metal oxide in a proportion of preferably 10 to 60 wt. %, more preferably 20 to 40 wt. %, although depending on the strength of the magnetic force. When containing the superparamagnetic metal oxide in a proportion within the specified range, the magnetic silica particulate carrier can be readily separated using a commercially available magnet.
For use in the invention, a particulate carrier having the following properties is most preferred:
(1) being magnetic silica particles containing a superparamagnetic iron oxide,
(2) having an outer surface area of at least 5 m2/g,
(3) comprising composites of fine particles each comprising a superparamagnetic metal oxide covered with silica, the composites having porous surfaces,
(4) having a superparamagnetic iron oxide content of 10 to 60 wt. %,
(5) having a specific surface area of 5 to 800 m2/g,
(6) having a pore diameter of 50 to 500 nm,
(7) having a pore volume of 200 to 5000 mm3/g, and
(8) having a particle diameter of 0.5 to 15.0 xcexcm.
Process for Production of Particulate Carrier
The particulate carrier for use in the invention can be produced by a known process disclosed in, for example, Japanese Unexamined Patent Publication No. 47273/1994.
For example, a magnetic silica particulate carrier containing an iron oxide can be produced by the following process. First, triiron tetraoxide is added to an alcohol solution of tetraethoxysilane, and dispersed and wetted using an ultrasonic dispersion apparatus. A catalyst for hydrolyzing tetraethoxysilane is added to the dispersion and ultrasonically dispersed so as to deposit silica on the particle surfaces of triiron tetraoxide. Sodium silicate is added to the resulting dispersion, and an organic solvent and a surfactant (a toluene solution of sorbitan monostearate) are added to form a w/o emulsion. The emulsion is added to an aqueous solution of ammonium sulfate, followed by thorough stirring. The resulting mixture was subjected to filtration separation, washing with water, alcohol precipitation and drying, to thereby obtain desired spherical silica particles.
Nucleic Acid Extraction Method Using Nucleic Acid-binding Particulate Carrier
In the method of the invention, the nucleic acid-binding particulate carrier having the above properties is used to extract nucleic acids from a material containing nucleic acids. Specifically, nucleic acids are extracted by contacting a nucleic acid-binding particulate carrier having the above properties with a material containing nucleic acids so that the nucleic acids are bound and adsorbed to the carrier.
More specifically, according to the method of the invention, nucleic acids are extracted and isolated by the steps of:
(a) mixing a material containing nucleic acids, a nucleic acid-binding particulate carrier having a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g, and a nucleic acid extraction solution for allowing the nucleic acids to adsorb to the carrier, to thereby bind the nucleic acids to the carrier (adsorption step);
(b) separating a composite of the nucleic acids and the particulate carrier from the mixture obtained in Step (a) to remove contaminants (separation step); and
(c) eluting and collecting the nucleic acids from the composite (elution step).
Step (a)
The material containing nucleic acids for use in the invention may be a biological material, specifically, an animal-derived biological material such as blood (including whole blood, blood serum, blood plasma, etc.), urine, saliva or like body fluid, or a material derived from organisms other than animals, such as plants and microorganisms. The material for use in the invention may also be cells separated from the above organisms, cultures of said cells, or partially purified nucleic acids.
According to the present invention, the nucleic acids may be DNA or RNA. DNA may be double-stranded DNA, single-stranded DNA, plasmid DNA, genomic DNA, cDNA, etc. RNA may be RNA derived from exogenous parasites such as virus, bacteria and fungi, or endogenous RNA derived from organisms that produce these biological materials. Thus, RNA may be tRNA, mRNA, rRNA, etc.
The nucleic acid extraction solution for use in the invention is a solution capable of destroying nucleic acid-containing cells in the biological material to thereby expose the nucleic acids and allow the nucleic acids to bind to the particulate carrier. Preferred examples of such solutions include those containing substances capable of increasing hydrophobicity of the surfaces of glass or slica particles, such as chaotropic substances.
Specific examples of chaotropic substances include guanidine salt, potassium iodide, sodium iodide, (iso)thiocyanate sodium perchlorate and urea, and more specifically, guanidine thiocyanate, guanidine hydrochloride, sodium iodide, potassium iodide, sodium perchlorate and urea. Among these chaotropic substances, guanidine thiocyanate and guanidine hydrochloride can be preferably used since they have high inhibitory activity against ribonucleases that decompose RNA. Thus, the nucleic acid extraction solution is preferably an aqueous solution of a compound having inhibitory activity against nucleases, such as guanidine thiocyanate and/or guanidine hydrochloride. The concentration of the chaotropic substance in the nucleic acid extraction solution is usually 1.0 to 8.0 M, preferably 4.0 to 7.0 M, although depending on the kind of the chaotropic substance used. For example, guanidine hydrochloride, when employed, is used preferably at a concentration of 4.0 to 7.5 M. When employing guanidine thiocyanate, it is used preferably at a concentration of 3.0 to 5.5 M.
The nucleic acid extraction solution for use in the invention preferably contains a buffer. The buffer may be added to the extraction solution before use, or may be added as a buffer solution to the solution after dissolution of cells. Conventional buffers can be used in the invention without limitation. Preferred are those showing buffer action at neutral pH values, i.e., pH 5.0 to 9.0. Useful examples include Tris-hydrochloride, sodium tetraborate-hydrochloric acid, potassium dihydrogenphosphate-sodium tetraborate. Preferably, the buffer is used at a concentration of 1 to 500 mM in the nucleic acid extraction solution, and has a pH value of 6.0 to 9.0.
The nucleic acid extraction solution may contain a surfactant to destroy cell membranes or denature proteins in cells. Surfactants conventionally used for nucleic acid extraction from cells or the like may be employed without limitation. Specific examples include nonionic surfactants such as Triton surfactants and Tween surfactants, and anionic surfactants such as N-lauroyl sarcosine sodium. According to the invention, it is particularly preferred to use a nonionic surfactant at a concentration of 0.1 to 2.0 wt. % in the nucleic acid extraction solution. A anionic surfactant, when employed, is preferably used at a concentration similar to that of the nonionic surfactant.
Further, it is desirable that the nucleic acid extraction solution contains a suitable amount of a reducing agent such as 2-mercaptoethanol or dithiothreitol, in order to denature and inactivate proteins, in particular ribonucleases, contained in the sample.
The particulate carrier is usually used as suspended in sterilized water, a 1 mM to 5 M NaCl solution, a 1 mM to 5 M LiCl solution or like solution.
According to the invention, relative to 0.01 to 2 ml of a sample containing nucleic acids, the particulate carrier is used usually 0.5 to 10 mg, preferably 2 to 5 mg, and the nucleic acid extraction solution is used usually 0.09 to 9 ml, preferably 0.9 to 2 ml, although depending on the nucleic acid concentration of the sample. It should be noted, however, that the amounts of the particulate carrier and the nucleic acid extraction solution are not limited to the above ranges.
In the invention, the particulate carrier and a material containing nucleic acids are mixed with the nucleic acid extraction solution so that the nucleic acids are contacted with and adsorbed to the nucleic acid-binding particulate carrier, giving a composite of the particulate carrier and the nucleic acids.
To achieve thorough mixing, stirring can be carried out by vortex action, end-over-end action or magnetic action, for about 1 to 60 minutes. By stirring, nucleic acids, small amounts of proteins and saccharides in the sample are adsorbed to the nucleic acid-binding particulate carrier. The temperature for stirring is not limited.
Step (b)
The mixture obtained in Step (a) is separated into a solution containing contaminants unbound to the particulate carrier and the particulate carrier having nucleic acids bound thereto.
The separation can be carried out by centrifugation. Alternatively, when a magnetic particulate carrier is used, magnetic separation using a magnet or the like can be carried out with ease. The solution containing contaminants are removed by the separation.
After removing the solution containing contaminants, the particulate carrier may be washed with a washing solution as required, to elute unnecessary substances such as proteins, saccharides and lipids.
The washing solution is not limited, as long as it does not cause elution of nucleic acids from the particulate carrier and is capable of eluting proteins, saccharides and lipids from the carrier. Preferably, the carrier is washed with a solution containing a chaotropic substance and/or an alcohol solution. As a chaotropic substance, at least one compound selected from the group consisting of guanidine thiocyanate, guanidine hydrochloride and sodium thiocyanate is preferred. More preferred is a 4.0 to 7.5 M guanidine hydrochloride solution. A preferred alcohol solution is a 40 to 100%, preferably 60 to 100% alcohol solution. Usable alcohols include ethanol, propanol, isopropanol and butanol, among which ethanol is preferred. In particular, it is desirable to wash the carrier with a solution containing a chaotropic substance, preferably with a solution similar to the nucleic acid extraction solution used in Step (a), since said extraction solution is effective for removing lipids, saccharides and proteins, and then it is desirable to wash the carrier with a 40 to 100% ethanol solution. When employing alcohol solutions, use of two alcohol solutions with different concentrations is more effective. For example, the carrier can be washed with a 70% ethanol solution and then with a 99% ethanol solution. The washing is carried out once or twice.
As required, the carrier with bound nucleic acids is dried by, for example, heating.
Step (c)
Nucleic acids are eluted using a nucleic acid eluate, from the particulate carrier obtained in Step (b), which has nucleic acids bound (adsorbed) thereto.
The nucleic acid eluate for use in the invention is not limited as long as it is capable of eluting nucleic acids from the carrier. Preferred examples include water and TE buffer solution (10 mM Tris-hydrochloride, 1.0 mM EDTA; pH 8.0).
Specifically, the nucleic acid eluate is mixed with the carrier having the nucleic acids, and the mixture is allowed to stand or stirred, to elute the nucleic acids from the carrier. The reaction temperature is about 10 to 50xc2x0 C. and the reaction time is about 1 to 60 minutes. The amount of the eluate to be added is about 0.01 to 0.2 ml, preferably about 0.05 to 0.15 ml, per 1 to 50 mg of the particulate carrier.
Thereafter, the nucleic acid-containing solution is isolated from the carrier by centrifugation, or by magnetic separation using a magnet or the like when a magnetic particulate carrier is used.
Nucleic Acid Detection Method
The nucleic acid solution obtained in Step (c) need not be desalted as by dialysis or ethanol precipitation, or condensed, and can be subjected directly to enzyme reaction using a restriction enzyme, a DNA polymerase or the like. The target nucleic acid can be amplified by polymerase chain reaction (PCR) or nucleic acid sequence based amplification (NASBA) before detection by, for example, nucleic acid hybridization assay, using nucleic acid probes.
PCR can be carried out by, for example, the method described in Science 230:1350-1354, 1985. NASBA can be carried out by, for example, the method described in Nature 350:91-92, 1991, and Japanese Patents Nos. 2648802 and 2650159.
Nucleic acid hybridization assay can be carried out by the method described in, for example, Unexamined Japanese Patent Publication No. 189794/1994.
Nucleic Acid Extraction Kit
The nucleic acid extraction kit of the invention comprises at least the nucleic acid-binding particulate carrier, the nucleic acid extraction solution and the nucleic acid eluate. More specifically, the extraction kit of the invention comprises at least (1) a nucleic acid-binding particulate carrier having a particle diameter of 0.5 to 15.0 xcexcm, a pore diameter of 50 to 500 nm and a pore volume of 200 to 5000 mm3/g, (2) the nucleic acid extraction solution for allowing nucleic acids to adsorb to the carrier, and (3) the nucleic acid eluate for eluting nucleic acids. The amounts of the components (1) to (3) can be selected according to the purpose of use, and may be, for example, about 1 to 200 xcexcl of the component (1) (the concentration of the carrier being about 0.01 to 1 g/ml), about 0.1 to 10.0 ml of the component (2) and about 10 to 500 xcexcl of the component (3).

Method of extracting nucleic acids using particulate carrier


The disclosure provides for a compressed mode transmission gap in wireless communications. A user equipment (UE) may receiving a downlink dedicated physical channel (DPCH) having a slot-format and a spreading factor during a first compression interval. The downlink DPCH may not include a compressed-mode transmission gap during the first compression interval. The UE may receive the downlink DPCH having the same slot-format and the same spreading factor during a second compression interval. The UE may determine that the downlink DPCH includes a compressed-mode transmission gap during the second compression interval. A set of slots of the downlink DPCH during the transmission gap may be punctured. The UE may decode the downlink DPCH for the second compression interval based on a set of remaining slots received during the second compression interval. In an aspect, the UE may estimate a SIR based on a TPC command in a last slot of the transmission gap.

Field
Aspects of the present disclosure relate generally to wireless communication systems, and more particularly, to physical layer configuration of wireless communications.
Background
Wireless communication networks are widely deployed to provide various communication services such as telephony, video, data, messaging, broadcasts, and so on. Such networks, which are usually multiple access networks, support communications for multiple users by sharing the available network resources. One example of such a network is the UMTS Terrestrial Radio Access Network (UTRAN). The UTRAN is the radio access network (RAN) defined as a part of the Universal Mobile Telecommunications System (UMTS), a third generation (3G) mobile phone technology supported by the 3rd Generation Partnership Project (3GPP). The UMTS, which is the successor to Global System for Mobile Communications (GSM) technologies, currently supports various air interface standards, such as Wideband-Code Division Multiple Access (W-CDMA), Time Division-Code Division Multiple Access (TD-CDMA), and Time Division-Synchronous Code Division Multiple Access (TD-SCDMA). The UMTS also supports enhanced 3G data communications protocols, such as High Speed Packet Access (HSPA), which provides higher data transfer speeds and capacity to associated UMTS networks.
As the demand for mobile broadband access continues to increase, research and development continue to advance the UMTS technologies not only to meet the growing demand for mobile broadband access, but to advance and enhance the user experience with mobile communications.
Release-99 is a standard that defined dedicated channels between a user equipment (UE) and a base station or node B based on orthogonal variable spreading factor (OVSF) codes. Some wireless networks continue to rely on Release-99 channels for various services. For example, a voice call may be carried over a Release-99 dedicated channel (DCH). Release-99 channels allow for a compressed mode (CM) operation to allow a UE to perform measurements on another frequency or another radio access technology (RAT). The Release-99 CM operation may reduce a spreading factor for a DCH. Such spreading factor reduction may present difficulties for network management of a code tree or may result in additional interference when non-orthogonal codes are used. In view of the foregoing, it may be understood that alternative methods of compressed mode operation are desirable.