The present invention is directed to an intimal hypertrophy inhibitor containing as the active ingredient an oxyindole derivative represented by the following formula (I) or a salt thereof: ##STR1## (wherein R.sup.1 represents a hydrogen atom; a phenyl group which may be substituted by a lower alkyl group, a lower alkoxy group, a lower alkylaminoalkoxy group, a hydroxy group, an amino group, a lower alkylamino group, or a halogen atom; or a pyridyl group which may be substituted by a lower alkyl group, a lower alkoxy group, a lower alkylaminoalkoxy group, a hydroxy group, an amino group, a lower alkylamino group, a halogen atom, a lower alkoxycarbonyl group, or a carboxyl group; R.sup.2 represents a phenyl group which may be substituted; or a pyridyl group which may be substituted by a lower alkyl group, a lower alkoxy group, a lower alkylaminoalkoxy group, a hydroxy group, an amino group, a lower alkylamino group, a halogen atom, a lower alkoxycarbonyl group, or a carboxyl group; R.sup.3 represents a hydrogen atom; a lower alkyl, benzyl, or a benzenesulfonyl group which may be substituted; or an acyl group; R.sup.4 represents a hydrogen atom; a lower alkoxy group, a halogen atom, an amino group, a lower alkylamino group, a carboxyl group, a lower alkoxycarbonyl group; a phenylcarbomoyl group which may be substituted; or a trifluoromethyl group; X represents CH or N; n represents a number between 0 and 4 inclusive that indicates the number of substituents; and the broken/solid double line denotes a single bond or a double bond). The intimal hypertrophy inhibitor of the present invention exhibits excellent inhibitory action against intimal hypertrophy, and thus is useful as a preventive/therapeutic/ameliorating agent for proliferative vascular diseases such as restenosis after PTCA, arteriosclerosis, peripheral embolism, and angiitis.

Coronary arteriosclerosis is known to precede and be the major cause of the onset of various pathological conditions such as angina pectoris and myocardial infarction. Luminal narrowing caused by arteriosclerosis and loss of vascular elasticity bring about deficiency of nutrition and oxygen in cardiac muscular tissue, to thereby induce the aforementioned pathological conditions. Narrowing of vascular lumina is considered to be primarily caused by accumulation of foamy macrophages and cholesterol on the inner wall, and in addition, by cell-fibrous intimal hypertrophy caused by migration of smooth muscle cells of the media into the intima, and proliferation of the cells in the intima. In the treatment of angina pectoris and myocardial infarction, antithrombotic agents, vasodilator agents, etc. have been used for the principal purpose of ameliorating the symptoms. However, these agents have failed to serve as a radical remedy for the narrowing and loss of elasticity of vascular lumina caused by arteriosclerosis. Therefore, pharmaceuticals that are capable of preventing or treating intimal hypertrophy which causes angiostenosis are earnestly desired.
In recent years, angiostenosis has been surgically treated by percutaneous transluminal coronary angioplasty (hereinafter referred to as PTCA). PTCA is a therapy in which a balloon catheter is inserted by remote operation into the narrow segment through, for example, the femoral artery, without performance of thoracotomy, and the balloon is inflated in situ, to thereby physically achieve vasodilation. Due to advancement in manipulation technique of PTCA, more than 90% of PTCA cases show amelioration of symptoms immediately after PTCA. In addition, PTCA rarely involves death or signs of adverse side effects such as induction of myocardial infarction. Accordingly, PTCA is accepted as an excellent therapy. However, about 30-40% of the cases that undergo PTCA revert to restenosis at the same site, and if restenosis occurs, PTCA must be performed again, or alternatively, an aorta bypass-forming operation must be performed. This constitutes the most significant problem in the clinical field. Autopsy of cases in which death was caused by reblockage after PTCA revealed that intimal hypertrophy had occurred at the site at which the vascular cavity had been dilated, thus reblocking the site [see, for example, British Heart Journal, 58, 635-643 (1987), Human Pathology, 477-485 (1989)].
Therefore, it is considered that inhibition of intimal hypertrophy would be effective for the prevention of restenosis after PTCA and further for the treatment of arteriosclerosis.
The pharmaceuticals that were expected to open the way up to the remedy of the aforementioned diseases have been studied in both preclinical and clinical stages [American Heart Journal, 122, 171-187 (1991)]. The candidate pharmaceuticals were anticoagulants such as heparin; platelet aggregation inhibitors such as aspirin, dipyridamole, ticlopidine, prostacycline, and their derivatives; thromboxane A2 inhibitors such as trapidil; cell proliferation inhibitors such as ketanserin; calcium antagonists such as diltiazem and nifedipine; lipid decreasing agents such as fish oil, eicosapentaenoic acid and lovastatin; and anti-inflammatory agents such as steroids. However, after actual investigation in clinical situations, none of these drugs were found to have clear utility.
Tranilast is an intimal hypertrophy inhibitor considered to be developed to the most advanced level (Japanese Patent Application Laid-Open (kokai) No. 6-135829). However, due to its weak activity, this drug is not a satisfactory intimal hypertrophy inhibitor.
Thus, presently there are no effective drugs against intimal hypertrophy, and therefore, clinically useful pharmaceuticals are strongly called for.
The active ingredients of the present invention, oxindole derivatives, partially comprise known compounds. The known compounds include synthesis intermediates disclosed in Japanese Patent Publication (kokoku) No. 43-3195, Hungarian Patent Application Laid-Open No. 65452, U.S. Pat. No. 4,002,749, Chem. Ber. 91, 2095 and 91, 1898 (1958), and U.S. Pat. No. 3,413,299; UV absorbers disclosed in U.S. Pat. No. 3,428,649; antidepressant agents or tranquilizers disclosed in Japanese Patent Application Laid-Open (kokai) No. 47-8628; antinootropic agents disclosed in WO 91/01306; drugs for the treatment of central nervous diseases or stomach ulcers disclosed in WO 92/07830; and drugs for the treatment of asthma, rheumatic arthritis, and allergic rhinitis disclosed in WO 95/14667. In addition, myocardial infarction augmenting effect of these compounds is disclosed in European Journal of Medicinal Chemistry, 25(2), 187 (1990), ibid. 27(2), 167 (1992), and ibid. 28, 653 (1993). However, these compounds have not yet been known to serve as intimal hypertrophy inhibitors. Also, analogs of the active ingredients of the present invention, i.e., analogs of the oxindole derivatives, are disclosed in Japanese Patent Application Laid-Open (kokai) Nos. 62-29570, 6-501494, and 7-108900, among which Japanese Patent Application Laid-Open (kokai) Nos. 62-29570 and 6-501494 disclose tyrosine kinase inhibitory activity and Japanese Patent Application Laid-Open (kokai) No. 7-108900 discloses antioxidative action. However, intimal hypertrophy inhibitory action is not at all described in those publications.
Accordingly, the object of the present invention is to provide an excellent intimal hypertrophy inhibiting agent.


A non-visible radiation imaging system is provided in which an image is obtained based on non-visible infrared radiation of a subject. The image can be enhanced to increase its resolution. Additionally, the image can be combined with another image based on visible light for the subject. The system also provides for a step by step procedure which must be following to obtain a non-visible radiation image accurately for medical diagnostics. Further, a non-visible radiation diagnostic examination system and method are provided that perform an automatic diagnostic information extraction based on examination of the subject using one or more of the images.

Numerous imaging devices exist for generating images of human subjects based on electromagnetic radiation in the spectral band. However, additional information on one or more attributes of many subjects can be obtained from imaging electromagnetic radiation having wavelengths that fall above and/or below visible light in the infrared spectral band. For example, infrared light (e.g., thermal radiation) can be imaged to determine the temperature characteristics of the subject. Other radiation spectra having non-visible wavelengths that may provide useful information include ultraviolet light, X-rays, radio waves, and the like.
To date, three major technologies are used to detect and/or measure infrared light. A bolometer, which includes an extremely fine wire in an electrical circuit, can measure temperature based on a change in conductance in the wire. When only a particular spectrum of radiation, such as infrared light, is permitted to reach the bolometer, the radiation can be measured. Similarly, a pyroelectric device can measure radiation by exposing a particular type of crystal to a particular spectrum of radiation. Finally, a thermopile, which includes numerous thermocouple elements, can measure radiation based on temperature changes for each element.
Each technology has been used to create an imaging device for infrared light. In particular, a two-dimensional matrix of a selected detection technology can be combined with proper optics to generate a two-dimensional image of radiation in the infrared spectrum. A similar matrix design is used in modern digital cameras for generating visible light images. However, to date, infrared imaging solutions have lagged in both resolution and cost as compared to visible light imaging solutions. As a result, only small resolution infrared imaging systems that use a thermopile array are available for an affordable price (e.g., less than a few hundred dollars). However, these imaging systems have an insufficient resolution for many demanding medical applications.
To date, numerous solutions have been proposed that seek to obtain additional resolution from low-resolution images, particularly visible light images. These solutions include a one-pass super-resolution solution, a simple cubic or bilinear resampling, and the like. A more complex solution comprises a longer-term super-resolution approach that attempts to extract data through complex averaging methods from multiple low-resolution images.
Additional information on a subject can also be obtained by fusing two or more images. Image fusion combines images from one or more sensing modalities, e.g., infrared light and visible light, into a single presentation that retains the useful and unique information from both modalities. Properly done, an image fusion presentation can be synergistic. That is, the fused presentation allows the viewer to comprehend more of the totality of the subject being imaged. As a result, a need exists for an improved imaging and/or examination solution that can incorporate imaging of non-visible radiation (e.g., infrared light) in a cost-effective manner. In particular, a need exists for an imaging system and an examination system and method that generate and use an image based on non-visible radiation of the subject.
However, users must observe strict regime in order to acquire useful infrared images. Unless a strict process and procedure is followed, the infrared images will not be as useful in medical diagnostics on human beings or other living bodies. Proper environment control temperature, humidity control, emissivity verification, etc. are just some of the steps needed to be observed before a scientifically useful medical thermal image can be obtained.
Also, the infrared images must be processed by using innovative algorithms to enhance them so that most of the information from the infrared images, fused with visible images or as is infrared images, can be clarified, enhanced, or improved to obtain underlying human medical diagnostic information. It should be noted that typical infrared images obtainable from lower cost, affordable imagers are typically noisy and not very clear, which is not suitable or required for medical diagnostics and medical work.
Last but not the least, the medical infrared images must be processed in a systematic way without taking a lot of time from the healthcare provider to allow the provider with an easy and automated means to extract medical diagnostic information. Especially, when a provider with limited time is trying to arrive at a diagnostic conclusion, lots of images must be compared automatically to reduce human error and improve decision time simultaneously.


A terahertz continuous wave generator includes: an optical intensity modulator configured to modulate an optical signal into DSB optical signals; a local oscillator configured to generate a modulation signal for modulating the optical signal inputted to the optical intensity modulator into DSB optical signals; a notch filter configured to filter an optical signal with a specific frequency; an optical fiber amplifier configured to amplify an output signal of the optical intensity modulator; an optical circulator configured to transmit the optical signal inputted to the optical fiber amplifier to the notch filter and transmit the optical signal reflected from the notch filter to an input of the optical intensity modulator; an optical coupler configured to apply the optical signal to the optical intensity modulator; and an OE converter configured to photomix the DSB signals outputted through the notch filter.

Exemplary embodiments of the present invention relate to a terahertz continuous wave generator, and more particularly, to a terahertz continuous wave generator which generates a terahertz continuous wave (CW) through photomixing.
Recently, research has been actively conducted on photomixing that uses an optical signal to generate a CW signal having a frequency of nm-wave bands or more, which is not sensitive to low phase noise, temperature, and an operation environment such as a radio environment.
Photomixing is to generate a CW in a ultra-high frequency band, a mm-wave band, and a terahertz (THz) band, which has low phase noise and narrow FWHM (full wave half maximum), by beating two different optical signals through an opto-electric converter (OE converter).
Meanwhile, a terahertz continuous wave generator using photomixing generates two optical signals maintaining a strong correlation and having different wavelengths from one optical source and beats the generated signals, in order to minimize frequency drift and generate a stable CW THz signal.
The terahertz continuous wave generation system, which includes one optical source, passive optical parts, and an OE converter, generates optical signals having two or more kinds of wavelengths from the optical source, and then uses the generated optical signals. Therefore, since the correlation between two wavelengths to be beaten is high, the system may reduce phase noise and increase the frequency stability of the generated signals.
A method for generating a signal of a THz-range frequency from one optical source includes a mode locking laser method, a dual mode laser method, an injection locking method, double sideband-suppressed carrier (DSB-SC) generation, and a frequency comb method.
In the case of the mode locking laser method and the dual mode laser method, an optical device is difficult to manufacture, and the competitiveness thereof is insufficient in terms of the development cost and price of devices. Therefore, the effectiveness of commercialization for the mode locking laser method and the dual mode laser method becomes a problem. In the injection locking method, a locking process should be performed to obtain a desired frequency signal. Therefore, since the operation condition is complicated, the injection locking method has a limit in effectiveness. The frequency comb method includes an optical source, an optical intensity modulator, two optical phase modulators, arrayed waveguide grating (AWG), and an OE converter. In the frequency comb method, the frequency of a generated signal is variable depending on the performance of the optical phase modulator and the AWG, and two expensive optical phase modulators and expensive AWG are required. The DSB-SC generation is a kind of optical heterodyne method and has a simple configuration. Furthermore, it is easy to obtain a desired frequency signal, compared with the above-described methods. Therefore, much research has been conducted on the DSB-SC generation.
A terahertz continuous wave generator using the DSB-SC generation includes one laser diode (LD), passive optical parts, an optical fiber amplifier and so on. The passive optical parts include an optical intensity modulator, an ultrahigh frequency local oscillator (LO), an OE converter, an optical filter and so on, and the optical fiber amplifier is configured to amplify generated DSB signals.
The laser diode is an optical carrier generation source for generating two optical signals having different wavelengths, between which the correlation is high, in the terahertz continuous wave generator using the DSB-SC generation. Such an optical carrier is modulated into DSB signals by a local oscillator signal which is outputted from the ultrahigh frequency local oscillator and then inputted to the optical intensity modulator. The DSB-modulated optical signals are inputted to the OE converter to generate a THz CW signal through photomixing.
In general, an optical filter such as a notch filter or AWG is used to remove an optical carrier existing with the DSB signals on an optical spectrum, in order to increase the stability of the generated THz CW signal and reduce noise. Furthermore, an optical amplifier such as an Er-doped fiber amplifier is used to compensate for attenuation of the DSB signals caused by an insertion loss of the optical filter and increase the magnitude of the DSB signals, thereby generating a THz CW with high power.
The above-described configuration is a related art for helping an understanding of the present invention, and does not mean a related art which is widely known in the technical field to which the present invention belongs.
The conventional terahertz continuous wave generator using the DSB-SC generation may reduce phase noise and increase the frequency stability of a generated signal. However, since an expensive optical source for generating an optical carrier should be used, a manufacturing cost inevitably increases.

Electronics and Telecommunications Research Institute


A process and apparatus for enhancing in-tube heat transfer within a tubular member. A fluid is introduced into the tubular member and then directed in a downstream direction through a coil of the tubular member. The first coil defines a first coil axis. The fluid is further directed in the downstream direction through at least one downstream coil. Each downstream coil has a coil axis which is rotated at a coil switching angle, with respect to the coil axis of the immediately upstream coil. The fluid is maintained at a turbulent flow level within at least a portion of a coiled section of the tubular member. The fluid is then discharged from the tubular member.

1. Field of the Invention
This invention relates to a process and apparatus for achieving chaotic mixing of fluid flow through a coiled section of a tubular member for enhancing in-tube heat transfer, wherein the fluid flow is directed through a helical arrangement including coils having their corresponding coil axes positioned at some angle with respect to each other.
2. Description of Prior Art
Hydrodynamics of chaotic particle paths have been discussed within various publications, such as: Jones, Scott W., et al., Chaotic Advection by Laminar Flow in a Twisted Pipe, Journal of Fluid Mechanics, Vol. 209, p. 335 (1989), which states that chaotic advection of a passive scalar in steady laminar flow through a sequence of pipe bends leads to enhanced transverse and longitudinal stirrings, which is important to applications such as heat and mass transfer in piping systems; and Peerhossaini, H., et al., Chaotic Motion in the Dean Instability Flow--A Heat Exchanger Design, Bulletin of the American Physical Society, Program of the 43rd Annual Meeting of the Division of Fluid Dynamics, Vol. 35, No. 10, p. 2229 (1990). In Order Breaking in Dean Flow, Phys. Fluids A, Vol. 3, No. 5, p. 1029 (May 1991), Le Guer, Yves, et al. discuss combining large vortical structures in the flow with the dynamical behavior of open systems to enhance heat transfer and mixing, particularly in an apparatus having a succession of curved channels in which the curvature plane rotates 90.degree. between each of two adjacent curved elements. Although such reference suggests that chaotic mixing is expected to enhance the advection of passive scalars and therefore improve the efficiency of heat transfer from the walls, such reference only discusses heat transfer with respect to laminar flow and does not even suggest the impact of chaotic mixing in a turbulent flow regime.
Other references have discussed increasing in-tube heat transfer by various mechanical methods. For example, U.S. Pat. No. 2,115,769 teaches a radiant heating tube which has spiral grooves or corrugations along which burning gases are whirled and eddied. The centrifugal forces cause the burning gases to swirl as they pass through the conduit.
U.S. Pat. No. 4,444,357 teaches a method and apparatus for mixing two input streams at different temperatures to form an intermediate combined stream at a regulated temperature. A heat exchange coil acts as a quick-response, continuous-flow heat exchanger, which alternately extracts and returns heat from differing-temperature pulses as the same merge and blend through their travel within the coil.
U.S. Pat. No. 4,469,446 discloses a fluid handling apparatus which has a plurality of hollow airfoil shaped vanes which are positioned within a first gaseous stream, and a second gaseous stream is conveyed through the hollow vane and is discharged into the first stream through a slot at the trailing edge of the vane.
Conventional passive techniques for enhancement of in-tube conventional heat transfer can be roughly classified into two categories: (1) increased effective heat transfer area; and (2) increased mixing methods. External and internal fins fall into the former category of conventional art, while the latter category of conventional art includes increased turbulence intensity and increased unsteady flow through inserts of various kinds, regular mixing in recirculation zones and resonant enhancement of flow instabilities. Some devices, of course, do both.
It is apparent that conventional technology and prior art references consider only chaotic fluid flow as it relates to flow within the laminar regime. It is very likely that persons skilled in the art of fluid mechanics and heat and mass transfer have not considered the effects of chaotic mixing of flow within a turbulent regime, since it would be expected that the increased energy requirements for pressure drops associated with turbulent flow would far outweigh any energy benefits realized through increased heat exchange efficiency.

Process and apparatus for enhancing in-tube heat transfer by chaotic mixing


A storage container incorporates a housing defining a cavity therein. At least one support rail extends from the housing within the cavity having a plurality of encoded first moieties. At least one refrigerant brick having mating encoded second moieties configured to be received into the encoded first moieties couples the refrigerant brick within the cavity.

1. Field
Embodiments of the disclosure relate generally to the field of food service carts for airline in-flight food service and more particularly to a refrigerant brick and support rail interlocking configuration for multiple spaced refrigerant bricks carried intermediate food trays for distributed cooling.
2. Background
Galley carts are used on an aircraft to store food and beverages that need to be refrigerated and/or frozen during the duration of a flight. Conventional galley carts require the use of a powered heat exchanger (i.e., a chiller) to cool a housing cavity of the cart where the food products are stored. However, the heat exchanger discharges heated air to the environment surrounding the cart and consumes power that can be otherwise be used by the aircraft. Further, additional cabin cooling is needed to counteract the heated air discharged from the heat exchanger. As such, at least some known galley carts use dry ice to cool the housing cavity. However, when the dry ice is placed within the cart, it cools locally and the cooling stratifies along the height of the cart. To alleviate this problem, fans are added to the carts to circulate the cool air within the housing cavity. However, the fan requires a power source. Further, the dry ice releases CO2 gas as it sublimates. The CO2 can accumulate in the cabin and cause the CO2 concentration to increase above a predetermined threshold for CO2 in the cabin. Moreover, the cost of dry ice is increasing and airlines need to manage their dry ice supplies. The proposed invention allows airlines to stop using dry ice.
It is therefore desirable to provide a galley cart with distributed cooling which overcomes the limitations of current galley carts.

Insulated galley cart and method of using frozen refrigerant bricks to maximize storage volume and equalize temperature distribution


A method for chemical mechanical polishing (CMP) of a shallow trench isolation (STI) structure employs a sequence of slurry polishes. In the first step the substrate is polished with either silica-based slurry or diluted ceria-based slurry. The first polishing is at a higher removal rate than the second polishing step. The polishing proceeds with some planarization but does not expose the polish stop layer. After partial planarization, the high selectivity slurry was used to complete the process. Improved throughput, lower defects and good within wafer uniformity are achieved.

1. Field of Invention
This invention is directed to a method of chemical mechanical polishing and more particularly for planarizing shallow trench isolation or similar structures for integrated circuits.
2. Description of Related Art
Shallow trench isolation (STI) structures are formed between field effect transistors in an integrated circuit to prevent carriers such as electrons from drifting between adjacent device elements. The fabrication of STI structures involves photolithography to form the trench structures followed by filling of the trenches with insulators. Planarization of the filled trenches is usually accomplished by chemical mechanical polishing (CMP). The CMP process uses chemical etchants and abrasives in the form of aqueous slurries to polish the preliminary STI structures. The CMP process is not without throughput issues with larger diameter wafers or defect concerns such as shallow dishes from overpolishing, microscratches or across the wafer non-uniformity. The importance of overcoming some of the deficiencies or cost factors is evidenced by the technology developments directed to the subject as noted in the patent literature.
U.S. Pat. No. 6,365,520 (Rhoades et al) comprises CMP slurry for planarization of STI based on a mixture of two ranges of particle sizes.
U.S. Pat. No. 6,261,158 (Holland et al ) describes a two step CMP process for planarizing metal interconnects with the imposition of a cleansing/neutralization step as intermediate between stages of CMP.
U.S. Pat. No. 6,234,877 (Koos et al) provides a method of controlling the pH of the slurry composition by an intermediate cleansing rinse of the polishing pads with a diluting solution or a buffered solution.
U.S. Pat. No. 6,207,535 (Lee et al) teaches the thermal hardening of a portion of a third oxide layer prior to CMP of that layer in a STI fabrication process.
U.S. Pat. No. 6,190,999 (Hung et al) relates to a STI structure formed by a sequence of film depositions and the intermediate removal of the silicon nitride hard masking layer to expose the pad oxide layer. An insulating layer is formed over the trench; the pad oxide layer then acts as a polishing stop for planarizing the insulating layer.
U.S. Pat. No. 6,143,662 (Rhoades et al) describes for STI planarization a method of using a CMP slurry mixture of abrasive particles having a mean diameter of between 2 and 30 nm and larger abrasive particles having a mean diameter between 2 and 10 times the mean diameter of the small abrasive particles.
U.S. Pat. No. 6,117,748 (Lou et al) describes a dishing-free process for STI consisting of a two-step CMP with an oxide slurry followed by a poly slurry that stops at the surface of the nitride layer.
U.S. Pat. No. 5,652,177 (Pan) encompasses the sequences of forming a field isolation region by depositing an insulating layer, a polysilicon layer, and a nitride layer over a substrate followed by the lithographic patterning and etching of the silicon and nitride layers over the insulating layer. The nitride layer is removed prior to CMP and the polysilicon layer acts as a polishing stop.

Method for chemical mechanical polishing of a shallow trench isolation structure

Taiwan Semiconductor Manufacturing Company, Ltd.


The present invention provides ungulate animals, tissue and organs as well as cells and cell lines derived from such animals, tissue and organs, which lack expression of functional endogenous immunoglobulin loci. The present invention also provides ungulate animals, tissue and organs as well as cells and cell lines derived from such animals, tissue and organs, which express xenogenous, such as human, immunoglobulin loci. The present invention further provides ungulate, such as porcine genomic DNA sequence of porcine heavy and light chain immunogobulins. Such animals, tissues, organs and cells can be used in research and medical therapy. In addition, methods are provided to prepare such animals, organs, tissues, and cells.

An antigen is an agent or substance that can be recognized by the body as 'foreign'. Often it is only one relatively small chemical group of a larger foreign substance which acts as the antigen, for example a component of the cell wall of a bacterium. Most antigens are proteins, though carbohydrates can act as weak antigens. Bacteria, viruses and other microorganisms commonly contain many antigens, as do pollens, dust mites, molds, foods, and other substances. The body reacts to antigens by making antibodies. Antibodies (also called immunoglobulins (Igs)) are proteins that are manufactured by cells of the immune system that bind to an antigen or foreign protein. Antibodies circulate in the serum of blood to detect foreign antigens and constitute the gamma globulin part of the blood proteins. These antibodies interact chemically with the antigen in a highly specific manner, like two pieces of a jigsaw puzzle, forming an antigen/antibody complex, or immune complex. This binding neutralizes or brings about the destruction of the antigen.
When a vertebrate first encounters an antigen, it exhibits a primary humoral immune response. If the animal encounters the same antigen after a few days the immune response is more rapid and has a greater magnitude. The initial encounter causes specific immune cell (B-cell) clones to proliferate and differentiate. The progeny lymphocytes include not only effector cells (antibody producing cells) but also clones of memory cells, which retain the capacity to produce both effector and memory cells upon subsequent stimulation by the original antigen. The effector cells live for only a few days. The memory cells live for a lifetime and can be reactivated by a second stimulation with the same antigen. Thus, when an antigen is encountered a second time, its memory cells quickly produce effector cells which rapidly produce massive quantities of antibodies.
By exploiting the unique ability of antibodies to interact with antigens in a highly specific manner, antibodies have been developed as molecules that can be manufactured and used for both diagnostic and therapeutic applications. Because of their unique ability to bind to antigenic epitopes, polyclonal and monoclonal antibodies can be used to identify molecules carrying that epitope or can be directed, by themselves or in conjunction with another moiety, to a specific site for diagnosis or therapy. Polyclonal and monoclonal antibodies can be generated against practically any pathogen or biological target. The term polyclonal antibody refers to immune sera that usually contain pathogen-specific antibodies of various isotypes and specificities. In contrast, monoclonal antibodies consist of a single immunoglobulin type, representing one isotype with one specificity.
In 1890, Shibasaburo Kitazato and Emil Behring conducted the fundamental experiment that demonstrated immunity can be transmitted from one animal to another by transferring the serum from an immune animal to a non-immune animal. This landmark experiment laid the foundation for the introduction of passive immunization into clinical practice. However, wide scale serum therapy was largely abandoned in the 1940s because of the toxicity associated with the administration of heterologous sera and the introduction of effective antimicrobial chemotherapy. Currently, such polyclonal antibody therapy is indicated to treat infectious diseases in relatively few situations, such as replacement therapy in immunoglobulin-deficient patients, post-exposure prophylaxis against several viruses (e.g., rabies, measles, hepatitis A and B, varicella), and toxin neutralization (diphtheria, tetanus, and botulism). Despite the limited use of serum therapy, in the United States, more than 16 metric tons of human antibodies are required each year for intravenous antibody therapy. Comparable levels of use exist in the economies of most highly industrialized countries, and the demand can be expected to grow rapidly in developing countries. Currently, human antibody for passive immunization is obtained from the pooled serum of donors. Thus, there is an inherent limitation in the amount of human antibody available for therapeutic and prophylactic therapies.
The use of antibodies for passive immunization against biological warfare agents represents a very promising defense strategy. The final line of defense against such agents is the immune system of the exposed individual. Current defense strategies against biological weapons include such measures as enhanced epidemiologic surveillance, vaccination, and use of antimicrobial agents. Since the potential threat of biological warfare and bioterrorism is inversely proportional to the number of immune persons in the targeted population, biological agents are potential weapons only against populations with a substantial proportion of susceptible persons.
Vaccination can reduce the susceptibility of a population against specific threats; provided that a safe vaccine exists that can induce a protective response. Unfortunately, inducing a protective response by vaccination may take longer than the time between exposure and onset of disease. Moreover, many vaccines require multiple doses to achieve a protective immune response, which would limit their usefulness in an emergency to provide rapid prophylaxis after an attack. In addition, not all vaccine recipients mount a protective response, even after receiving the recommended immunization schedule.
Drugs can provide protection when administered after exposure to certain agents, but none are available against many potential agents of biological warfare. Currently, no small-molecule drugs are available that prevent disease following exposure to preformed toxins. The only currently available intervention that could provide a state of immediate immunity is passive immunization with protective antibody (Arturo Casadevall "Passive Antibody Administration (Immediate Immunity) as a Specific Defense Against Biological Weapons" from Emerging Infectious Diseases, Posted Sep. 12, 2002).
In addition to providing protective immunity, modern antibody-based therapies constitute a potentially useful option against newly emergent pathogenic bacteria, fungi, virus and parasites (A. Casadevall and M. D. Scharff, Clinical Infectious Diseases 1995; 150). Therapies of patients with malignancies and cancer (C. Botti et al, Leukemia 1997; Suppl 2:S55-59; B. Bodey, S. E. Siegel, and H. E. Kaiser, Anticancer Res 1996; 16(2):661), therapy of steroid resistant rejection of transplanted organs as well as autoimmune diseases can also be achieved through the use of monoclonal or polyclonal antibody preparations (N. Bonnefoy-Berard and J. P. Revillard, J Heart Lung Transplant 1996; 15(5):435-442; C. Colby, et al Ann Pharmacother 1996; 30(10):1164-1174; M. J. Dugan, et al, Ann Hematol 1997; 75(1-2):41 2; W. Cendrowski, Boll Ist Sieroter Milan 1997; 58(4):339-343; L. K. Kastrukoff, et al Can J Neurol Sci 1978; 5(2):175178; J. E. Walker et al J Neurol Sci 1976; 29(2-4):303309).
Recent advances in the technology of antibody production provide the means to generate human antibody reagents, while avoiding the toxicities associated with human serum therapy. The advantages of antibody-based therapies include versatility, low toxicity, pathogen specificity, enhancement of immune function, and favorable pharmacokinetics.
The clinical use of monoclonal antibody therapeutics has just recently emerged. Monoclonal antibodies have now been approved as therapies in transplantation, cancer, infectious disease, cardiovascular disease and inflammation. In many more monoclonal antibodies are in late stage clinical trials to treat a broad range of disease indications. As a result, monoclonal antibodies represent one of the largest classes of drugs currently in development.
Despite the recent popularity of monoclonal antibodies as therapeutics, there are some obstacles for their use. For example, many therapeutic applications for monoclonal antibodies require repeated administrations, especially for chronic diseases such as autoimmunity or cancer. Because mice are convenient for immunization and recognize most human antigens as foreign, monoclonal antibodies against human targets with therapeutic potential have typically been of murine origin. However, murine monoclonal antibodies have inherent disadvantages as human therapeutics. For example, they require more frequent dosing to maintain a therapeutic level of monoclonal antibodies because of a shorter circulating half-life in humans than human antibodies. More critically, repeated administration of murine immunoglobulin creates the likelihood that the human immune system will recognize the mouse protein as foreign, generating a human anti-mouse antibody response, which can cause a severe allergic reaction. This possibility of reduced efficacy and safety has lead to the development of a number of technologies for reducing the immunogenicity of murine monoclonal antibodies.
Polyclonal antibodies are highly potent against multiple antigenic targets. They have the unique ability to target and kill a plurality of "evolving targets" linked with complex diseases. Also, of all drug classes, polyclonals have the highest probability of retaining activity in the event of antigen mutation. In addition, while monoclonals have limited therapeutic activity against infectious agents, polyclonals can both neutralize toxins and direct immune responses to eliminate pathogens, as well as biological warfare agents.
The development of polyclonal and monoclonal antibody production platforms to meet future demand for production capacity represents a promising area that is currently the subject of much research. One especially promising strategy is the introduction of human immunoglobulin genes into mice or large domestic animals. An extension of this technology would include inactivation of their endogenous immunoglobulin genes. Large animals, such as sheep, pigs and cattle, are all currently used in the production of plasma derived products, such as hyperimmune serum and clotting factors, for human use. This would support the use of human polyclonal antibodies from such species on the grounds of safety and ethics. Each of these species naturally produces considerable quantities of antibody in both serum and milk.
Arrangement of Genes Encoding Immunoglobulins
Antibody molecules are assembled from combinations of variable gene elements, and the possibilities resulting from combining the many variable gene elements in the germline enable the host to synthesize antibodies to an extraordinarily large number of antigens. Each antibody molecule consists of two classes of polypeptide chains, light (L) chains (that can be either kappa (κ) L-chain or lambda (λ) L-chain) and heavy (H) chains. The heavy and light chains join together to define a binding region for the epitope. A single antibody molecule has two identical copies of the L chain and two of the H chain. Each of the chains is comprised of a variable region (V) and a constant region (C). The variable region constitutes the antigen-binding site of the molecule. To achieve diverse antigen recognition, the DNA that encodes the variable region undergoes gene rearrangement. The constant region amino acid sequence is specific for a particular isotype of the antibody, as well as the host which produces the antibody, and thus does not undergo rearrangement.
The mechanism of DNA rearrangement is similar for the variable region of both the heavy- and light-chain loci, although only one joining event is needed to generate a light-chain gene whereas two are needed to generate a complete heavy-chain gene. The most common mode of rearrangement involves the looping-out and deletion of the DNA between two gene segments. This occurs when the coding sequences of the two gene segments are in the same orientation in the DNA. A second mode of recombination can occur between two gene segments that have opposite transcriptional orientations. This mode of recombination is less common, although such rearrangements can account for up to half of all Vκ to Jκ joins; the transcriptional orientation of half of the human Vκ gene segments is opposite to that of the Jκ gene segments.
The DNA sequence encoding a complete V region is generated by the somatic recombination of separate gene segments. The V region, or V domain, of an immunoglobulin heavy or light chain is encoded by more than one gene segment. For the light chain, the V domain is encoded by two separate DNA segments. The first segment encodes the first 95-101 amino acids of the light chain and is termed a V gene segment because it encodes most of the V domain. The second segment encodes the remainder of the V domain (up to 13 amino acids) and is termed a joining or J gene segment. The joining of a V and a J gene segment creates a continuous exon that encodes the whole of the light-chain V region. To make a complete immunoglobulin light-chain messenger RNA, the V-region exon is joined to the C-region sequence by RNA splicing after transcription.
A heavy-chain V region is encoded in three gene segments. In addition to the V and J gene segments (denoted VH and JH to distinguish them from the light-chain VL and JL), there is a third gene segment called the diversity or DH gene segment, which lies between the VH and JH gene segments. The process of recombination that generates a complete heavy-chain V region occurs in two separate stages. In the first, a DH gene segment is joined to a JH gene segment; then a VH gene segment rearranges to DJH to make a complete VH-region exon. As with the light-chain genes, RNA splicing joins the assembled V-region sequence to the neighboring C-region gene.
Diversification of the antibody repertoire occurs in two stages: primarily by rearrangement ("V(D)J recombination") of Ig V, D and J gene segments in precursor B cells resident in the bone marrow, and then by somatic mutation and class switch recombination of these rearranged Ig genes when mature B cells are activated. Immunoglobulin somatic mutation and class switching are central to the maturation of the immune response and the generation of a "memory" response.
The genomic loci of antibodies are very large and they are located on different chromosomes. The immunoglobulin gene segments are organized into three clusters or genetic loci: the κ, λ, and heavy-chain loci. Each is organized slightly differently. For example, in humans, immunoglobulin genes are organized as follows. The λ light-chain locus is located on chromosome 22 and a cluster of Vλ gene segments is followed by four sets of Jλ gene segments each linked to a single Cλ gene. The κ light-chain locus is on chromosome 2 and the cluster of Vκ, gene segments is followed by a cluster of Jκ gene segments, and then by a single Cκ gene. The organization of the heavy-chain locus, on chromosome 14, resembles that of the κ locus, with separate clusters of VH, DH, and JH gene segments and of CH genes. The heavy-chain locus differs in one important way: instead of a single C-region, it contains a series of C regions arrayed one after the other, each of which corresponds to a different isotype. There are five immunoglobulin heavy chain isotypes: IgM, IgG, IgA, IgE and IgD. Generally, a cell expresses only one at a time, beginning with IgM. The expression of other isotypes, such as IgG, can occur through isotype switching.
The joining of various V, D and J genes is an entirely random event that results in approximately 50,000 different possible combinations for VDJ(H) and approximately 1,000 for VJ(L). Subsequent random pairing of H and L chains brings the total number of antibody specificities to about 107 possibilities. Diversity is further increased by the imprecise joining of different genetic segments. Rearrangements occur on both DNA strands, but only one strand is transcribed (due to allelic exclusion). Only one rearrangement occurs in the life of a B cell because of irreversible deletions in DNA. Consequently, each mature B cell maintains one immunologic specificity and is maintained in the progeny or clone. This constitutes the molecular basis of the clonal selection; i.e., each antigenic determinant triggers the response of the pre-existing clone of B lymphocytes bearing the specific receptor molecule. The primary repertoire of B cells, which is established by V(D)J recombination, is primarily controlled by two closely linked genes, recombination activating gene (RAG)-1 and RAG-2.
Over the last decade, considerable diversity among vertebrates in both Ig gene diversity and antibody repertoire development has been revealed. Rodents and humans have five heavy chain classes, IgM, IgD, IgG, IgE and IgA, and each have four subclasses of IgG and one or two subclasses of IgA, while rabbits have a single IgG heavy chain gene but 13 genes for different IgA subclasses (Burnett, R. C et al. EMBO J. 8:4047; Honjo, In Honjo, T, Alt. F. W. T. H. eds, Immunoglobulin Genes p. 123 Academic Press, New York). Swine have at least six IgG subclasses (Kacskovics, I et al. 1994 J Immunol 153:3565), but no IgD (Butler et al. 1996 Inter. Immunol 8:1897-1904). A gene encoding IgD has only been described in rodents and primates. Diversity in the mechanism of repertoire development is exemplified by contrasting the pattern seen in rodents and primates with that reported for chickens, rabbits, swine and the domesticated Bovidae. Whereas the former group have a large number of VH genes belonging to seven to 10 families (Rathbun, G. In Hongo, T. Alt. F. W. and Rabbitts, T. H., eds, Immunoglobulin Genes, p. 63, Academic press New York), the VH genes of each member of the latter group belong to a single VH gene family (Sun, J. et al. 1994 J. Immunol. 1553:56118; Dufour, V et al. 1996, J Immunol. 156:2163). With the exception of the rabbit, this family is composed of less than 25 genes. Whereas rodents and primates can utilize four to six JH segments, only a single JH is available for repertoire development in the chicken (Reynaud et al. 1989 Adv. Immunol. 57:353). Similarly, Butler et al. (1996 Inter. Immunol 8:1897-1904) hypothesized that swine may resemble the chicken in having only a single JH gene. These species generally have fewer V, D and J genes; in the pig and cow a single VH gene family exists, consisting of less than 20 gene segments (Butler et al, Advances in Swine in Biomedical Research, eds: Tumbleson and Schook, 1996; Sinclair et al, J. Immunol. 159: 3883, 1997). Together with lower numbers of J and D gene segments, this results in significantly less diversity being generated by gene rearrangement. However, there does appear to be greater numbers of light chain genes in these species. Similar to humans and mice, these species express a single κ light chain but multiple λ light chain genes. However, these do not seem to affect the restricted diversity that is achieved by rearrangement.
Since combinatorial joining of more than 100 VH, 20-30 DH and four to six JH gene segments is a major mechanism of generating the antibody repertoire in humans, species with fewer VH, DH or JH segments must either generate a smaller repertoire or use alternative mechanisms for repertoire development. Ruminants, pigs, rabbits and chickens, utilize several mechanisms to generate antibody diversity. In these species there appears to be an important secondary repertoire development, which occurs in highly specialized lymphoid tissue such as ileal Peyer's patches (Binns and Licence, Adv. Exp. Med. Biol. 186: 661, 1985). Secondary repertoire development occurs in these species by a process of somatic mutation which is a random and not fully understood process. The mechanism for this repertoire diversification appears to be templated mutation, or gene conversion (Sun et al, J. Immunol. 153: 5618, 1994) and somatic hypermutation.
Gene conversion is important for antibody diversification in some higher vertebrates, such as chickens, rabbits and cows. In mice, however, conversion events appear to be infrequent among endogenous antibody genes. Gene conversion is a distinct diversifying mechanism characterized by transfers of homologous sequences from a donor antibody V gene segment to an acceptor V gene segment. If donor and acceptor segments have numerous sequence differences then gene conversion can introduce a set of sequence changes into a V region by a single event. Depending on the species, gene conversion events can occur before and/or after antigen exposure during B cell differentiation (Tsai et al. International Immunology, Vol. 14, No. 1, 55-64, January 2002).
Somatic hypermutation achieves diversification of antibody genes in all higher vertebrate species. It is typified by the introduction of single point mutations into antibody V(D)J segments. Generally, hypermutation appears to be activated in B cells by antigenic stimulation.
Production of Animals with Humanized Immune Systems
In order to reduce the immunogenicity of antibodies generated in mice for human therapeutics, various attempts have been made to replace murine protein sequences with human protein sequences in a process now known as humanization. Transgenic mice have been constructed which have had their own immunoglobulin genes functionally replaced with human immunoglobulin genes so that they produce human antibodies upon immunization. Elimination of mouse antibody production was achieved by inactivation of mouse Ig genes in embryonic stem (ES) cells by using gene-targeting technology to delete crucial cis-acting sequences involved in the process of mouse Ig gene rearrangement and expression. B cell development in these mutant mice could be restored by the introduction of megabase-sized YACs containing a human germline-configuration H- and κ L-chain minilocus transgene. The expression of fully human antibody in these transgenic mice was predominant, at a level of several 100 μg/l of blood. This level of expression is several hundred-fold higher than that detected in wild-type mice expressing the human Ig gene, indicating the importance of inactivating the endogenous mouse Ig genes in order to enhance human antibody production by mice.
The first humanization attempts utilized molecular biology techniques to construct recombinant antibodies. For example, the complementarity determining regions (CDR) from a mouse antibody specific for a hapten were grafted onto a human antibody framework, effecting a CDR replacement. The new antibody retained the binding specificity conveyed by the CDR sequences (P. T. Jones et al. Nature 321: 522-525 (1986)). The next level of humanization involved combining an entire mouse VH region with a human constant region such as gamma1 (S. L. Morrison et al., Proc. Natl. Acad. Sci., 81, pp. 6851-6855 (1984)). However, these chimeric antibodies, which still contain greater than 30% xenogeneic sequences, are sometimes only marginally less immunogenic than totally xenogeneic antibodies (M. Bruggemann et al., J. Exp. Med., 170, pp. 2153-2157 (1989)).
Subsequently, attempts were carried out to introduce human immunoglobulin genes into the mouse, thus creating transgenic mice capable of responding to antigens with antibodies having human sequences (Bruggemann et al. Proc. Nat'l. Acad. Sci. USA 86:6709-6713 (1989)). Due to the large size of human immunoglobulin genomic loci, these attempts were thought to be limited by the amount of DNA, which could be stably maintained by available cloning vehicles. As a result, many investigators concentrated on producing mini-loci containing limited numbers of V region genes and having altered spatial distances between genes as compared to the natural or germline configuration (See, for example, U.S. Pat. No. 5,569,825). These studies indicated that producing human sequence antibodies in mice was possible, but serious obstacles remained regarding obtaining sufficient diversity of binding specificities and effector functions (isotypes) from these transgenic animals to meet the growing demand for antibody therapeutics.
In order to provide additional diversity, work has been conducted to add large germline fragments of the human Ig locus into transgenic mammals. For example, a majority of the human V, D, and J region genes arranged with the same spacing found in the unrearranged germline of the human genome and the human Cμ and Cδ constant regions was introduced into mice using yeast artificial chromosome (YAC) cloning vectors (See, for example, WO 94/02602). A 22 kb DNA fragment comprising sequences encoding a human gamma-2 constant region and the upstream sequences required for class-switch recombination was latter appended to the foregoing transgene. In addition, a portion of a human kappa locus comprising Vκ, Jκ and Cκ region genes, also arranged with substantially the same spacing found in the unrearranged germline of the human genome, was introduced into mice using YACS. Gene targeting was used to inactivate the murine IgH & kappa light chain immunoglobulin gene loci and such knockout strains were bred with the above transgenic strains to generate a line of mice having the human V, D, J, Cμ, Cδ and Cγ2 constant regions as well as the human Vκ, Jκ and Cκ region genes all on an inactivated murine immunoglobulin background (See, for example, PCT patent application WO 94/02602 to Kucherlapati et al.; see also Mendez et al., Nature Genetics 15:146-156 (1997)).
Yeast artificial chromosomes as cloning vectors in combination with gene targeting of endogenous loci and breeding of transgenic mouse strains provided one solution to the problem of antibody diversity. Several advantages were obtained by this approach. One advantage was that YACs can be used to transfer hundreds of kilobases of DNA into a host cell. Therefore, use of YAC cloning vehicles allows inclusion of substantial portions of the entire human Ig heavy and light chain regions into a transgenic mouse thus approaching the level of potential diversity available in the human. Another advantage of this approach is that the large number of V genes has been shown to restore full B cell development in mice deficient in murine immunoglobulin production. This ensures that these reconstituted mice are provided with the requisite cells for mounting a robust human antibody response to any given immunogen. (See, for example, WO 94/02602; L. Green and A. Jakobovits, J. Exp. Med. 188:483-495 (1998)). A further advantage is that sequences can be deleted or inserted onto the YAC by utilizing high frequency homologous recombination in yeast. This provides for facile engineering of the YAC transgenes.
In addition, Green et al. Nature Genetics 7:13-21 (1994) describe the generation of YACs containing 245 kb and 190 kb-sized germline configuration fragments of the human heavy chain locus and kappa light chain locus, respectively, which contained core variable and constant region sequences. The work of Green et al. was recently extended to the introduction of greater than approximately 80% of the human antibody repertoire through introduction of megabase sized, germline configuration YAC fragments of the human heavy chain loci and kappa light chain loci, respectively, to produce XenoMouse™ mice. See, for example, Mendez et al. Nature Genetics 15:146-156 (1997), Green and Jakobovits J. Exp. Med. 188:483-495 (1998), European Patent No. EP 0 463 151 B1, PCT Publication Nos. WO 94/02602, WO 96/34096 and WO 98/24893.
Several strategies exist for the generation of mammals that produce human antibodies. In particular, there is the "minilocus" approach that is typified by work of GenPharm International, Inc. and the Medical Research Council, YAC introduction of large and substantially germline fragments of the Ig loci that is typified by work of Abgenix, Inc. (formerly Cell Genesys). The introduction of entire or substantially entire loci through the use microcell fusion as typified by work of Kirin Beer Kabushiki Kaisha.
In the minilocus approach, an exogenous Ig locus is mimicked through the inclusion of pieces (individual genes) from the Ig locus. Thus, one or more VH genes, one or more DH genes, one or more JH genes, a mu constant region, and a second constant region (such as a gamma constant region) are formed into a construct for insertion into an animal. See, for example, U.S. Pat. Nos. 5,545,807, 5,545,806, 5,625,825, 5,625,126, 5,633,425, 5,661,016, 5,770,429, 5,789,650, 5,814,318, 5,591,669, 5,612,205, 5,721,367, 5,789,215, 5,643,763; European Patent No. 0 546 073; PCT Publication Nos. WO 92/03918, WO 92/22645, WO 92/22647, WO 92/22670, WO 93/12227, WO 94/00569, WO 94/25585, WO 96/14436, WO 97/13852, and WO 98/24884; Taylor et al. Nucleic Acids Research 20:6287-6295 (1992), Chen et al. International Immunology 5:647-656 (1993), Tuaillon et al. J. Immunol. 154:6453-6465 (1995), Choi et al. Nature Genetics 4:117-123 (1993), Lonberg et al. Nature 368:856-859 (1994), Taylor et al. International Immunology 6:579-591 (1994), Tuaillon et al. J. Immunol. 154:6453-6465 (1995), and Fishwild et al. Nature Biotech. 14:845-851 (1996).
In the microcell fusion approach, portions or whole human chromosomes can be introduced into mice (see, for example, European Patent Application No. EP 0 843 961 A1). Mice generated using this approach and containing the human Ig heavy chain locus will generally possess more than one, and potentially all, of the human constant region genes. Such mice will produce, therefore, antibodies that bind to particular antigens having a number of different constant regions.
While mice remain the most developed animal for the expression of human immunoglobulins in humans, recent technological advances have allowed for progress to begin in applying these techniques to other animals, such as cows. The general approach in mice has been to genetically modify embryonic stem cells of mice to knock-out murine immunoglobulins and then insert YACs containing human immunoglobulins into the ES cells. However, ES cells are not available for cows or other large animals such as sheep and pigs. Thus, several fundamental developments had to occur before even the possibility existed to generate large animals with immunoglobulin genes knocked-out and that express human antibody. The alternative to ES cell manipulation to create genetically modified animals is cloning using somatic cells that have been genetically modified. Cloning using genetically modified somatic cells for nuclear transfer has only recently been accomplished.
Since the announcement of Dolly's (a cloned sheep) birth from an adult somatic cell in 1997 (Wilmut, I., et al (1997) Nature 385: 810-813), ungulates, including cattle (Cibelli, J et al 1998 Science 280: 1266-1258; Kubota, C. et al. 2000 Proc. Nat'l. Acad. Sci. 97: 990-995), goats (Baguisi, A. et al., (1999) Nat. Biotechnology 17: 456-461), and pigs (Polejaeva, I. A., et al. 2000 Nature 407: 86-90; Betthauser, J. et al. 2000 Nat. Biotechnology 18: 1055-1059) have been cloned.
The next technological advance was the development of the technique to genetically modify the cells prior to nuclear transfer to produce genetically modified animals. PCT publication No. WO 00/51424 to PPL Therapeutics describes the targeted genetic modification of somatic cells for nuclear transfer.
Subsequent to these fundamental developments, single and double allele knockouts of genes and the birth of live animals with these modifications have been reported. Between 2002 and 2004, three independent groups, Immerge Biotherapeutics, Inc. in collaboration with the University of Missouri (Lai et al. (Science (2002) 295: 1089-1092) & Kolber-Simonds et al. (PNAS. (2004) 101(19):7335-40)), Alexion Pharmaceuticals (Ramsoondar et al. (Biol Reprod (2003)69: 437-445) and Revivicor, Inc. (Dai et al. (Nature Biotechnology (2002) 20: 251-255) & Phelps et al. (Science (2003) January 17; 299(5605):411-4)) produced pigs that lacked one allele or both alleles of the alpha-1,3-GT gene via nuclear transfer from somatic cells with targeted genetic deletions. In 2003, Sedai et al. (Transplantation (2003) 76:900-902) reported the targeted disruption of one allele of the alpha-1,3-GT gene in cattle, followed by the successful nuclear transfer of the nucleus of the genetically modified cell and production of transgenic fetuses.
Thus, the feasibility of knocking-out immunoglobulin genes in large animals and inserting human immunoglobulin loci into their cells is just now beginning to be explored. However, due to the complexity and species differences of immunoglobulin genes, the genomic sequences and arrangement of Ig kappa, lambda and heavy chains remain poorly understood in most species. For example, in pigs, partial genomic sequence and organization has only been described for heavy chain constant alpha, heavy chain constant mu and heavy chain constant delta (Brown and Butler Mol Immunol. 1994 June; 31(8):633-42, Butler et al Vet Immunol Immunopathol. 1994 October; 43(1-3):5-12, and Zhao et al J Immunol. 2003 Aug. 1; 171(3):1312-8).
In cows, the immunoglobulin heavy chain locus has been mapped (Zhao et al. 2003 J. Biol. Chem. 278:35024-32) and the cDNA sequence for the bovine kappa gene is known (See, for example, U.S. Patent Publication No. 2003/0037347). Further, approximately 4.6 kb of the bovine mu heavy chain locus has been sequenced and transgenic calves with decreased expression of heavy chain immunoglobulins have been created by disrupting one or both alleles of the bovine mu heavy chain. In addition, a mammalian artificial chromosome (MAC) vector containing the entire unarranged sequences of the human Ig H-chain and κ L-chain has been introduced into cows (TC cows) with the technology of microcell-mediated chromosome transfer and nuclear transfer of bovine fetal fibroblast cells (see, for example, Kuroiwa et al. 2002 Nature Biotechnology 20:889, Kuroiwa et al. 2004 Nat Genet. June 6 Epub, U.S. Patent Publication Nos. 2003/0037347, 2003/0056237, 2004/0068760 and PCT Publication No. WO 02/07648).
While significant progress has been made in the production of bovine that express human immunoglobulin, little has been accomplished in other large animals, such as sheep, goats and pigs. Although cDNA sequence information for immunoglobulin genes of sheeps, goats and pigs is readily available in Genbank, the unique nature of immunoglobulin loci, which undergo massive rearrangements, creates the need to characterize beyond sequences known to be present in mRNAs (or cDNAs). Since immunoglobulin loci are modular and the coding regions are redundant, deletion of a known coding region does not ensure altered function of the locus. For example, if one were to delete the coding region of a heavy-chain variable region, the function of the locus would not be significantly altered because hundreds of other function variable genes remain in the locus. Therefore, one must first characterize the locus to identify a potential "Achilles heel".
Despite some advancements in expressing human antibodies in cattle, greater challenges remain for inactivation of the endogenous bovine Ig genes, increasing expression levels of the human antibodies and creating human antibody expression in other large animals, such as porcine, for which the sequence and arrangement of immunoglobulin genes are largely unknown.
It is therefore an object of the present invention to provide the arrangement of ungulate immunoglobin germline gene sequence.
It is another object of the present invention to provide novel ungulate immunoglobulin genomic sequences.
It is a further object of the present invention to provide cells, tissues and animals lacking at least one allele of a heavy and/or light chain immunoglobulin gene.
It is another object of the present invention to provide ungulates that express human immunoglobulins.
It is a still further object of the present invention to provide methods to generate cells, tissues and animals lacking at least one allele of novel ungulate immunoglobulin gene sequences and/or express human immunoglobulins.

Ungulates with genetically modified immune systems


A battery connection plate has a plurality of bus bars provided on a base plate section and is to be attached to a battery assembly containing a stacked plurality of batteries so that the bus bars are connected to terminals on the batteries. The base plate section has a wiring member attachment section to which a plurality of signal output wires connected to the bus bars can be wired, and window sections through which bosses can be inserted are formed on wiring paths of the signal output wires at the wiring member attachment section. The signal output wires are wired so as to bypass or wind around the bosses inserted into the window sections, and then the bosses are pulled out from the window sections, and thereby surplus length sections are formed on the signal output wires.

As eco-friendly vehicles, electric vehicles or hybrid cars are increasing.
Such a vehicle includes, mounted therein, a power source constituted by a battery assembly containing a stacked plurality of batteries.
That kind of power supply device contains a battery assembly configured such that batteries each having a positive electrode at one end and a negative electrode at the other end are alternately reversely coupled to one another. The power supply device further contains a battery connection plate configured such that a bus bar having two holes into which electrodes of adjacent batteries are inserted is provided on a base plate section made of a synthetic resin so as to form a connection section with respect to the batteries in order to serially connect the batteries.
As battery connection plates like the above, there is one in which a flexible section is formed, by each of a predetermined number of connection sections, on a slit provided on a base plate section with an escape hole therebetween in order to adjust positional deviation between an electrode and a hole of a bus bar, a base plate section is divided by a slit and both sides of each base plate section are coupled by a pair of hinge-like flexible sections, or pitch adjusting unit configured such that a base plate section is divided by each connection section and the connection sections are mutually coupled by a hinge-like flexible section, is provided (see, for example, Patent Document 1).


Embodiments of probes for selectively detecting compounds having a thiol group and an amino group, e.g., cysteine and/or homocysteine, are disclosed, along with methods and kits for detecting the compounds in neutral media with the probes. The probes have a structure according to the general formula where R1-R4 independently are hydrogen hydroxyl, halogen, thiol, thioether, lower aliphatic, or lower alkoxy, x is an integer from 0 to 4, and each R5 independently is halogen, hydroxyl, thiol, thioether, lower aliphatic, or lower alkoxy. Embodiments of the disclosed probes are capable of undergoing condensation/cyclization reactions with cysteine and/or homocysteine. Cysteine and/or homocysteine can be selectively detected and identified by determining fluorescence emission of the probes at characteristic wavelengths.

Biological thiols are essential in maintaining the appropriate redox status of proteins, cells and organisms (Wood et al., Trends Biochem. Sci. 2003, 28, 32-40; Schultz et al., Eur. J. Biochem. 2000, 267, 4904-4911). Cysteine (Cys) is an essential amino acid that is involved in protein synthesis, detoxification, and metabolism. Elevated levels of Cys have been associated with neurotoxicity (Wang et al., J. Neurosci. 2001, 21, 3322-3331). Cys deficiency is involved in slowed growth, hair depigmentation, edema, lethargy, liver damage, muscle and fat loss, skin lesions, and weakness (Shahrokhian, Anal. Chem. 2001, 73, 5972-5978). Homocysteine (Hcy) has been implicated in various types of vascular and renal diseases. Elevated Hcy (e.g., >12 μM) in blood is a well-known risk factor for cardiovascular, Alzheimer's disease, neutral tube defects, complications during pregnancy, inflammatory bowel disease, and osteoporosis (Seshadri et al., N. Eng. J. Med. 2002, 346, 476-483; Refsum et al., Annu. Rev. Med. 1998, 49, 31-62). Therefore, the determination of Cys and Hcy in vivo is correlated to physiological functions in diagnosing disease. However, because Cys and Hcy levels are associated with different diseases despite their similar structures, a need exists for a method to discriminate between Cys and Hcy.
Significant effort has gone into the development of colorimetric, phosphorescent, and fluorescent probes for these thiol-containing amino acids to achieve high sensitivity, low cost and ease of detection. To date, most of the indicators or dosimeters are based on the strong nucleophilicity of the thiol group, and various mechanisms have been employed, including Michael addition, cleavage reactions, and others. Though these probes show high sensitivity toward thiol-containing compounds, the direct detection of Cys (or Hcy) is hampered due to interference from other thiols.
For example, Cys and Hcy are known to undergo cyclization with aldehydes to form thiazolidines (or thiazinanes). Because both the sulfhydryl and the amino groups contribute to the cyclization, aldehyde cyclization enables selectivity for Cys and Hcy over other common thiols such as glutathione (GSH). However, since the aminothiol moieties of Cys and Hcy have similar reactivities towards aldehydes in general, discrimination of them from each other is challenging using heterocycle formation (Li et al., Chem. Comm. 2009, 5904-5906; Lee et al., Chem. Commun. 2008, 6173-6175; Tanaka et al., Chem. Commun. 2004, 1762-1763; Li et al., Chem. Commun. 2009, 5904-5906; Duan et al., Tetrahedron Lett. 2008, 49, 6624-6627; Kim et al., Tetrahedron Lett. 2008, 49, 4879-4881; Zhang et al., Tetrahedron Lett. 2007, 48, 2329-2333; Zhang et al., Org. Lett. 2009, 11, 1257-1260; Lim et al., Chem. Commun. 2010, 46, 5707-5709).

Fluorescence detection of cysteine and homocysteine


The object of the invention is a motor vehicle latch demonstrating a locking mechanism comprising a catch and at least a pawl, a latch plate (1), whereby the locking mechanism can be accommodated in the latch plate (1) and the latch plate (1) demonstrates an inlet area (12), whereby a latch holder (13) can be inserted to interact with the locking mechanism in the inlet area (12) and with at least a deformation (11) formed in a peripheral area of the inlet area (12) in the latch plate (1), whereby the deformation (11) is arranged at least in places directly on an edge of the inlet area (12).

Latches which retain the moving parts on the motor vehicle in their specified position are used to close motor vehicle doors, flaps or sliding doors, to name just a few examples. Normally, the latch systems or latches demonstrate a locking mechanism, consisting of a catch and at least a pawl which can be ratcheted into one another in order for example to encompass a latch holder attached to the motor vehicle chassis in order to thus, for example, retain a motor vehicle door in its position.
The locking mechanism, catch and pawl are pivotably accommodated on a latch plate, whereby the latch plate and the locking mechanism part axes incorporating the locking mechanism parts are made of metal, in particular steel. Due to the locking mechanism and the bearing parts of the latching system being made of steel, the latching system is capable of guaranteeing sufficient stability and thus safety even in extreme situations. Extreme situations for a latching system of a motor vehicle occur, for example, in the case of an accident, whereby the latching systems can be stressed beyond a multiple of the usual stresses.
In order to guarantee sufficient safety in these extreme situations too, tensile tests are performed in which the motor vehicle latches are subjected to a stress occurring in an accident. Under these simulated extreme conditions, the motor vehicle latches are subjected to tensile stress until failure of the locking mechanism. Failure means, for example, during tensile stress between the latch holder and the locking mechanism in the direction of the locking mechanism plane the latching unit comprising a motor vehicle latch and a latch holder are pulled apart until failure occurs of one of the components of the motor vehicle latch and/or the latch holder. To enable a latching unit to be used in the motor vehicle the latching unit must withstand tensile stress above the specifications of the car manufacturer before failure of one or several components of the latching unit occurs. In order to conform to the requirements of the motor vehicle manufacturer, different solutions have become known to stabilize a latch plate and/or a motor vehicle latch.
DE 20 2012 007 326 U1 discloses a motor vehicle latch with a latch case, furthermore with a locking mechanism accommodated in the latch case comprising a catch and a pawl and with an additional reinforcing element, whereby the reinforcing element is formed as a reinforcing plate connecting both rotational axes of both the catch and the pawl at a distance from the latch case. The latch case or the latch plate, it being pointed out that the latch case and latch plate can be used as synonyms, is U-shaped in the cross-section. As the legs of the U-shaped latch case are arranged fundamentally vertically to the motor vehicle transverse direction or y-direction, the specification of the latch case ensures that the rectangular supporting structure is arranged vertically to this y-direction. Any forces in this motor vehicle transverse direction are thus absorbed by the locking mechanism or the supporting structure and are introduced into this latch case. The latch case and reinforcing plate hereby from a storage cage for the locking mechanism. Consequently, deformation forces can be resiliently absorbed in the case of a crash.
From DE 10 2015 116 805 A1 a generic motor vehicle latch has become known demonstrating a locking mechanism comprising at least a catch and a pawl and a latch plate, whereby the catch and the pawl are pivotably accommodated on the latch plate and the latch plate demonstrates at least two apertures for mounting of the motor vehicle latch which demonstrates an inlet slot for a latch holder and whereby a splay opposite the inlet slot is provided for on the latch plate and the splay demonstrates at least a semi-circular end area opposite the inlet slot. The latch plate which is predominantly flat demonstrates deformations in the form of beads which extend around a recess, at least in places. The recess serves as an inlet area for a latch holder. The recess can also be described as an infeed section. In addition to the beads formed in the flat surface of the latch case, the latch plate demonstrates a splay arranged circumferentially at least in places which enables stabilization of the latch plate in the case of extreme stress.
A problem always posed in the further development of motor vehicle latches is that on the one hand safety which fulfills requirements needs to be guaranteed to enable the specified stress limitations to be attained. On the other hand, the automotive industry is striving to make motor vehicles lighter overall.


A resist composition which generates acid upon exposure and exhibits changed solubility in a developing solution by the action of acid, and which includes a base component which exhibits changed solubility in a developing solution by the action of acid, and a nitrogen-containing compound which has a boiling point of 50 to 200° C., a conjugate acid thereof having a pKa of 0 to 7, and a photodecomposable base; and a method of forming a resist pattern using the resist composition.

In lithography techniques, for example, a resist film composed of a resist material is formed on a substrate, and the resist film is subjected to selective exposure, followed by development, thereby forming a resist pattern having a predetermined shape on the resist film. A resist material in which the exposed portions of the resist film become soluble in a developing solution is called a positive-type, and a resist material in which the exposed portions of the resist film become insoluble in a developing solution is called a negative-type.
In recent years, in the production of semiconductor elements and liquid crystal display elements, advances in lithography techniques have lead to rapid progress in the field of pattern miniaturization. Typically, these miniaturization techniques involve shortening the wavelength (increasing the energy) of the exposure light source. Conventionally, ultraviolet radiation typified by g-line and i-line radiation has been used, but nowadays KrF excimer lasers and ArF excimer lasers are starting to be introduced in mass production of the semiconductor elements. Furthermore, research is also being conducted into lithography techniques that use an exposure light source having a wavelength shorter (energy higher) than these excimer lasers, such as electron beam (EB), extreme ultraviolet radiation (EUV), and X ray.
Resist materials for use with these types of exposure light sources require lithography properties such as a high resolution capable of reproducing patterns of minute dimensions, and a high level of sensitivity to these types of exposure light sources.
As a resist material that satisfies these conditions, a chemically amplified composition is used, which includes a base material component that exhibits a changed solubility in a developing solution under the action of acid and an acid generator component that generates acid upon exposure.
For example, in the case where the developing solution is an alkali developing solution (alkali developing process), a chemically amplified positive resist which contains, as a base component (base resin), a resin which exhibits increased solubility in an alkali developing solution under action of acid, and an acid generator is typically used. If the resist film formed using the resist composition is selectively exposed during formation of a resist pattern, then within the exposed portions, acid is generated from the acid generator component, and the action of this acid causes an increase in the polarity of the base resin, making the exposed portions soluble in the alkali developing solution. Thus, by conducting alkali developing, the unexposed portions remain to form a positive resist pattern.
On the other hand, in a case where such a base resin is applied to a solvent developing process using a developing solution containing an organic solvent (organic developing solution), when the polarity of the base resin is increased, the solubility of the exposed portions in an organic developing solution is relatively decreased. As a result, the unexposed portions of the resist film are dissolved and removed by the organic developing solution, and a negative resist pattern in which the exposed portions are remaining is formed. Such a solvent developing process for forming a negative-tone resist composition is sometimes referred to as "negative-tone developing process" (for example, see Patent Document 1).
In general, the base resin for a chemically amplified resist composition contains a plurality of structural units for improving lithography properties and the like. For example, in the case of a resin composition which exhibits increased solubility in an alkali developing solution by the action of acid, a structural unit containing an acid decomposable group which is decomposed by the action of acid generated from an acid generator component and exhibits increased polarity is used. Further, a structural unit containing a lactone-containing cyclic group or a structural unit containing a polar group such as a hydroxy group is used (for example, see Patent Document 2).
On the other hand, as acid generators usable in a chemically amplified resist composition, various types have been proposed including, for example, onium salt acid generators; oxime sulfonate acid generators; diazomethane acid generators; nitrobenzylsulfonate acid generators; iminosulfonate acid generators; and disulfone acid generators.
Further, as the miniaturization of patterns proceeds, a chemically amplified resist composition further containing an acid diffusion control agent, for obtaining an excellent contrast and for preventing roughness of a resist pattern and pattern collapse, as well as a base component and an acid generator component.
Patent Document 3 discloses a radiation-sensitive resin composition containing a compound which has a characteristic that, at unexposed portions of the resist film, an ion-exchange reaction with acid generated from an acid generator component and the like upon exposure is caused, thereby preventing diffusion of acid; and at exposed portions of the resist film, the compound is decomposed thereby, losing the basicity thereof with respect to acid generated from an acid generator component and the like.

Resist composition and method of forming resist pattern


The invention relates to an LED package having a metal reflective layer for focusing and emitting light through a side of the package, and a manufacturing method of the same. The LED package includes a substrate with an electrode formed thereon, a light emitting diode chip disposed on the substrate, and an encapsulant covering the LED chip and the substrate to protect the LED chip. The LED package also includes a metal reflective layer surrounding side surfaces of the encapsulant to form a light transmitting surface on a top surface of the encapsulant. The invention minimizes light loss, improves luminance, can be mass-produced as a PCB type, and adopts EMC transfer molding to minimize irregular color distribution, thereby improving optical quality.

1. Field of the Invention
The present invention relates to a Light Emitting Diode (LED) package having a metal reflective layer for focusing and emitting light through one side of the package, and a method of manufacturing the same. More particularly, the invention relates to an LED package with a metal reflective layer minimized in light loss and improved in luminance, which can be mass-manufactured as a PCB type in a small size unaffected by the size of an LED chip, and is improved in productivity.
2. Description of the Related Art
In general, amobile phone or PDA adopts a light emitting diode (LED) package in various sizes for a backlight.
As the backlights are becoming slimmer, the LED packages adopted therein are also becoming slimmer these days.
The LED package 300 shown in FIG. 1 is described in U.S. Patent No. 2003-0094622, in which a reflective layer 312 is formed of a lead frame for an LED chip 310 mounted in the package 300, and the package is sealed with a sealing plate 314 that covers the LED chip 310 and the reflective layer 312.
In this conventional LED package 300, the reflective layer 312 is made of an Ag-plated layer to focus and emit light from the LED chip 310 through one side of the package.
However, in such a conventional structure, the LED chip 310 is formed in a recess 322 of the substrate, and a separate sealing plate 314 covers the recess 322, and thus limiting automated manufacturing and mass-production.
FIGS. 2(a) and 2(b) illustrate a different structure of LED package 400 of the prior art. An LED chip 412 is mounted on a substrate 410, and a molded part 414 with a cavity therein is adhered on the substrate 410. Then, a resin solution with phosphor and epoxy mixed therein is injected into the space 414a to be cured, and the complete structure is diced. However, as an additional process is required to adhere the molded part 414 on the substrate 410, the manufacturing process is not efficient in terms of productivity.
In addition, as shown in FIG. 3, in the conventional LED package 400, the resin solution with the phosphor and epoxy mixed therein is injected into the space 414a in the molded part 414 and cured for about 1 hour to form an encapsulant 416. In such a curing process, the phosphor in the epoxy 418a of the encapsulant 416 tends to precipitate, causing irregular color distribution. Therefore, such a conventional LED package 400 yields low-uniformity and rather mediocre color development.

Light emitting diode package with metal reflective layer and method of manufacturing the same


Providing a zoom lens system having a high zoom ratio, a small diameter of the first lens group, capable of being effectively retracted upon retracting the lens barrel. The system includes, in order from an object, a first lens group having positive refractive power, a second lens group having negative refractive power, a third lens group having positive refractive power, a fourth lens group having positive refractive power, and a fifth lens group having positive refractive power. When zooming from a wide-angle end state to a telephoto end state, the first, second, third, and fourth lens groups move along an optical axis and the second lens group moves along a zoom trajectory being concave to the object. The fourth lens group is composed of a front lens group having positive power and a rear lens group having negative power with a space in between. Given conditional expressions are satisfied.

1. Field of the Invention
The present invention relates to a zoom lens system suitable for an electronic still camera.
2. Related Background Art
In a zoom lens system suitable for an electronic still camera, a zoom lens system whose first lens group is fixed, or a zoom lens system whose first lens group is movable has been known in Japanese Patent Application Laid-Open Nos. 6-281862 and 2003-177318.
However, in a zoom lens system whose first lens group is fixed, although the lens diameter of the first lens group can be small for focusing, it becomes difficult to make a zoom ratio large with securing high optical performance, so that the zoom ratio is about 5.7. Moreover, although the zoom ratio is tried to be made large abut from 7.6 to 9.7, aberration correction becomes insufficient in return for making the zoom ratio large, so that it becomes problem that optical performance deteriorates.
Moreover, in a zoom lens system whose first lens group is movable disclosed in Japanese Patent Application Laid-Open No. 6-281862, although optical performance is sufficient, the zoom ratio is small about from 1.9 to 3.7 and the half angle of view in the telephoto end state is about from 11.7° to 17.3° that is relatively large.
Furthermore, in a zoom lens system whose zoom ratio is made large by making the first lens group movable disclosed in Japanese Patent Application Laid-Open No. 2003-177318, although the zoom ratio is large about from 7.4 to 7.5 and the half angle of view in the telephoto end state is sufficiently small about from 4.6° to 4.7°, optical performance is not sufficient.


Compositions and methods are provided for depletion of pluripotent cells. In one embodiment of the invention, methods are provided for depletion of pluripotent cells from a mixed population of differentiated cells and stem cells, to provide a population of cells substantially free of pluripotent stem cells. Monoclonal antibodies useful in depletion and in identification of pluripotent stem cells are also provided.

Regenerative medicine is the process of creating living, functional tissues to repair or replace tissue or organ function lost due to age, disease, damage, or congenital defects. This field holds the promise of regenerating damaged tissues and organs in the body by introducing outside cells, tissue, or even whole organs to integrate and become a part of tissues or replace whole organ. Importantly, regenerative medicine has the potential to solve the problem of the shortage of organs available for donation compared to the number of patients that require life-saving organ transplantation.
One key to the success of regenerative medicine strategies has been the ability to isolate and generate stem cells, including pluripotent stem cells. In one aspect, pluripotent stem cells can be differentiated into a necessary cell type, where the mature cells are used to replace tissue that is damaged by disease or injury. This type of treatment could be used to replace neurons damaged by spinal cord injury, stroke, Alzheimer's disease, Parkinson's disease, or other neurological problems. Cells grown to produce insulin could treat people with diabetes and heart muscle cells could repair damage after a heart attack. This list could conceivably include any tissue that is injured or diseased.
The generation of pluripotent stem cells that are genetically identical to an individual provides unique opportunities for basic research and for potential immunologically-compatible novel cell-based therapies. Methods to reprogram primate somatic cells to a pluripotent state include differentiated somatic cell nuclear transfer, differentiated somatic cell fusion with pluripotent stem cells, and direct reprogramming to produce induced pluripotent stem cells (iPS cells) (Takahashi K, et al. (2007) Cell 131:861-872; Park I H, et al. (2008) Nature 451:141-146; Yu J, et al. (2007) Science 318:1917-1920; Kim D, et al. (2009) Cell Stem Cell 4:472-476; Soldner F, et al. (2009) Cell. 136:964-977; Huangfu D, et al. (2008) Nature Biotechnology 26:1269-1275; Li W, et al. (2009) Cell Stem Cell 4:16-19).
A significant first hurdle in stem cell-based therapy is the differentiation of pluripotent cells into a desired tissue type. Such methods currently rely on the step-wise introduction of factors and conditions to guide the cells down a developmental pathway, resulting eventually in a mature or committed progenitor cell that can transplanted into a patient.
The promise of pluripotent stem cells is that they can form any type of cell in the body. The trouble is that when implanted into an animal they do just that, forming all tissue types in the form of teratomas. These teratomas are one reason why it is necessary to mature the embryonic stem cells into highly purified adult cell types before they are considered appropriate for implanting into humans. The mature cells are restricted to their one identity and don't appear to revert to a teratoma-forming cell. However, even when researchers have learned to mature cells into a single cell type, getting those cells pure enough to eliminate the risk of remaining immature cells forming teratomas has been extremely difficult.
Clinical applications may utilize billions of cells injected into large numbers of clinically diverse patients, many of whom will be expected to live out their whole lifetimes (i.e. pediatric patients). As such it is important to thoroughly deplete residual pluripotent stem cells.
A caveat to the clinical application of ES/iPS derived derivatives is that the in vivo implantation of even trace amounts of undifferentiated cells may result in uncontrolled teratomas. The present invention addresses this issue.

Depletion of teratoma-forming pluripotent stem cells

THE BOARD OF TRUSTEES OF THE LELAND STANFORD JUNIOR UNIVERSITY


An automatic disk changer including: a pair of arms adapted to clamp a disk; a pair of coupling bars which are coupled through gear mechanisms to the base end portions of the arms, respectively; a cam adapted to swing the arms with the aid of the coupling bars; and a disk presence/absence detector which is provided confronted with one of the coupling bars. In the disk changer, the two arms are swung by the cam with the aid of the coupling bars, and when the angle formed by the two arms becomes smaller than the angle formed when the disk is actually clamped between them, the disk presence/absence detector is operated by the one of the coupling bar.

1. Field of the Invention
This invention relates to an automatic disk changer which picks up a desired one among a plurality of optical disks or magnet-optical disks such as compact disks (CDs) and mini-disks (MDs) which are arranged at predetermined intervals (hereinafter referred to merely as "disks", when applicable) so that data are recorded thereon or reproduced therefrom or erased therefrom.
2. Description of the Related Art
An example of an automatic disk changer of this type has been disclosed in Japanese Patent Unexamined Publication No. Sho. 61-156563. The automatic disk changer comprises a casing in which a disk arranging stand is provided on which a number of disks are arranged at predetermined intervals, and a carriage which is reciprocated along the disk arranging stand. The carriage has: a disk lifting mechanism which is adapted to slightly lift a specified disk among the disks arranged on the disk arranging stand; a loading mechanism which rolls the disk thus lifted to place it on the carriage; and a disk mounting mechanism which mounts the disk thus placed on a player body provided on the carriage.
The above-described automatic disk changer is disadvantageous in the following point: As was described above, the disk is delivered from the disk arranging stand to the disk mounting mechanism by means of the disk lifting mechanism and the loading mechanism; however, the disk delivering operation is carried out even if the specified disk is not present on the disk arranging stand, resulting in the occurrence of loss time.


A multi-pair cable having a plurality of twisted conductor pairs and a shielding arrangement. The shielding arrangement including at least one shielding component. The shielding component including a length of tape encased by a dielectric material.

A wide variety of cable arrangements having twisted conductor pairs are utilized in the telecommunications industry. In some cable arrangements, the twisted conductor pairs are separated by one or more filler components. In yet other arrangements, the cable includes shielding that surrounds the twisted conductor pairs, and the one or more filler components. The shielding reduces the occurrence of crosstalk between adjacent cables and thereby improves signal transmission performance of the twisted conductor pairs.
Cable shielding is commonly provided in the form of a conductive tape. The conductive tape surrounds the entire circumference of the cable core (i.e., the twisted conductor pairs, and the filler) to provide complete cable shielding. In particular, the conductive tape is wrapped around the entire cable core in an overlapping manner such that no gaps exist. Such shielded cables are expensive, typically require grounding, and further require specific connectors that accommodate the shielding.
In general, improvement has been sought with respect to existing cable assemblies, generally to reduce costs associated with twisted pair cables, and improve signal transmission performance of twisted pair cables.

Twisted pairs cable with shielding arrangement


Systems and methods for classifying a set of documents are provided. In some aspects, a method includes receiving a subset of the set of documents. The method also includes automatically determining at least one classification rule for the subset of documents based on the documents in the subset. At least a true-positive threshold proportion of documents in the subset of documents follows the at least one classification rule. At most a false-positive threshold proportion of documents in the set of documents and not in the subset of documents follow the at least one classification rule. The method also includes storing the at least one classification rule in association with the subset of documents.

Users of document management or electronic messaging services may receive many different documents or messages related to many different topics every day. To arrange their documents or messages and to be able to easily locate interesting or important documents or messages, users oftentimes apply labels to their documents or messages or place the documents or messages into folders. For example, in an electronic messaging service, a user may assign messages from members of his/her family the label "family," and messages concerning taxes, mortgages, or retirement the label "finance." One drawback of this labeling scheme is that users spend a lot of time reviewing their messages and determining that a particular message needs to be associated with a particular label or folder. Occasionally, users may forget to assign a label to a message, making locating and retrieving the message at a later time more difficult. As the foregoing illustrates, a technique for automatically classifying documents or messages by label or folder may be desirable.

Classifying documents based on automatically detected rules