Abstract: The present invention relates to compounds useful as inhibitors of protein kinases. The invention also provides pharmaceutically acceptable compositions comprising said compounds and methods of using the compositions in the treatment of various disease, conditions, or disorders.

Abstract: Methods and reagents are provided for determining endocytosis, using a cell expressing an externally polypeptide labeled cell membrane receptor, and as a reagent an antibody to said label conjugated to a fragment of an enzyme fragment complementation pair. Compounds are tested for their effect on endocytosis by complexing the reagent with said cells, adding the test compound at any time in relation to the complexing, allowing any endocytosis to occur, proteolytically degrading external enzyme fragment, adding protease inhibitor and the complementary member of the enzyme fragment complementation pair and substrate. The product provides a detectable signal related to the amount of endocytosis that occurred. The method is readily automated as all steps can occur in a single vessel without separations and washings.

Abstract: Methods are disclosed for producing electrochemiluminescence by electrochemically oxidizing an acridan compound at an electrode in the presence of a peroxide. Maintaining a sufficiently positive potential results in continuous oxidation of the acridan compound to an acridinium compound which reacts with peroxide to produce the luminescence. Light emission can be reversibly and repeatedly cycled on and off by sweeping the potential between two values. The acridan compounds can be provided with a labeling group for linking to an analyte or analyte binding partner. The present electrochemiluminescent reaction can find use in assay methods for detecting analytes by immunoassays and nucleic acid assays.

Abstract: This invention provides a method for determining the presence of hemolyzed erythrocytes in blood by detecting erythrocyte adenylate kinase in a serum sample from the blood. This invention also provides a method for diagnosing a hemolytic condition in a subject suspected to be suffering from hemolysis, as well as a method for monitoring hemolysis in a subject undergoing treatment for a hemolytic condition.

Abstract: Aqueous solutions of steroid compounds which have biological activity and have a tendency to oxidative degradation at temperatures between 2 and 8° C. on storage in excess of several months are stabilized by the addition of chelators. Aqueous solutions of particular interest are protein containing solutions which mimic the behavior of human bodily fluids such as serum and are therefore suitable as standards for immunoassays for steroids in such bodily fluids. Chelators of particular interest are transition metal chelators especially those which efficiently sequester iron.

Abstract: It has been found that casein and salts of casein are useful as replacements for, or in addition to, BSA as materials for coating solid phases, particularly magnetic particles, used for immunoassays and other binding assays for separation of the desired analyte. By using casein, immunoassays having improved stability and few discordant samples have been developed. Casein used at a concentration of 0.05-4.0 grams per gram of paramagnetic particle (optimally approximately 0.78-1.2 grams of casein per gram of magnetic particle) has been found to confer this benefit. In addition, a process for coating solid phases has been invented, said process comprising the mixing of casein with magnetic particles at 30-60° C. for 5-180 hours, said process resulting in casein-coated paramagnetic particles which either (1) already have combined therewith active ingredients needed in the binding assay or (2) are capable of reacting with active ingredients needed in the binding assay.

Abstract: A sensor for detecting an analyte in an environment includes a first reaction system including a first enzyme and a substrate for the first enzyme. The analyte inhibits the reaction of the substrate catalyzed by the first enzyme (in other words, the analyte inhibits the first enzyme). The sensor further includes at least a second reaction system that reacts to produce a first detectable state when the first enzyme is inhibited. In some embodiments, the reaction of the first reaction system can produce a second detectible state, different from the first detectible state. Another sensor for detecting an analyte in an environment includes a first reaction system including a first enzyme or a first substrate for the first enzyme. In this embodiment, the analyte is either a substrate for the first enzyme if the first reaction system includes the first enzyme or the first enzyme if the first reaction system includes the first substrate.

Abstract: The present invention discloses a one-pot, high-throughput method for the measurement of the amount of a nucleotide generated in a cell. The method is particularly effective in measuring changes in cyclic adenosine 3′,5′-monophosphate (cAMP) coupled to cell receptors in insects.

Abstract: This invention relates methods for conditioning affinity chromatography resins to decrease leaching of the ligand during purification. The methods involve incubating the resin in a buffered solution of a hydroxyalkylamine compound (e.g., ethanolamine) prior to use of the resin for an affinity purification. The treatment removes unstably bound ligand from the resin.

Abstract: A method of improving a phenotypic defect in a cell that contains a conformationally defective target protein wherein the conformational defect causes the phenotype defect, comprising contacting a first cell that expresses said conformationally defective target protein with an amount of a protein stabilizing agent that is effective to improve the conformational defect, thereby improving the phenotypic defect of the first cell in comparison with a second cell having the same conformationally defective target protein and phenotypic defect, wherein the second cell is not contacted with the protein stabilizing agent.

Abstract: In a competitive receptor binding assay for detecting TSH-receptor auto-antibodies in a biological sample, the sample is reacted in a reaction mixture which contains (i) a TSH-receptor or TSH-receptor preparation; (ii) a primary competitor, for example labelled TSH; and (iii) an agent for separating a complex composed of the TSH-receptor and the elements bound thereto of the reaction mixture from the liquid phase. According to the invention, the reaction is carried out in the presence of at least one monoclonal or polyclonal antibody specific against a partial peptide sequence of the TSH eceptor. This specific antibody is used to immobilize a complex of TSH-receptor and primary competitor and/or as secondary competitor for another part of the TSH-receptor auto-antibodies expected in a sample. The primary or secondary competitors are or can be selectively labelled.

Abstract: Methods of enhancing sensitivity and specificity of an assay measuring enzymatic activity in a sample by measuring enzymatic activity in the sample in the presence and absence of a specific inhibitor of the enzymatic activity are provided. Methods of measuring carboxypeptidase A levels and total carboxypeptidase A levels, wherein procarboxypeptidase A is converted to carboxypeptidase A by addition of clostripain, in a biological fluid with a carboxypeptidase A substrate, specificity of which is enhanced by addition of a carboxypeptidase A specific inhibitor are also provided. In addition, methods of diagnosing acute pancreatitis by measurement of carboxypeptidase A levels and pancreatic cancer by measurement of total carboxypeptidase A levels are also provided.

Abstract: It has been found that casein and salts of casein are useful as replacements for, or in addition to, BSA as materials for coating solid phases, particularly magnetic particles, used in immunoassays and other binding assays for separation of the desired analyte. By using casein, immunoassays having improved stability and fewer discordant samples have been developed. Casein used at a concentration of 0.05-4.0 grams per gram of paramagnetic particle (optimally approximately 0.78-1.2 grams of casein per gram of magnetic particle) has been found to confer this benefit.

Abstract: A reagent for a drug susceptibility testing which is simple and highly reproducible is provided. A microplate for a drug susceptibility testing wherein a drug and a color reagent containing a tetrazolium salt, 1-methoxy-5-methylphenazinium methylsulfate (1-methoxy PMS), potassium ferricyanide and potassium ferrocyanide are formed into a solid phase on the microplate by a vacuum drying, and a kit consisting of the microplate and a medium for a cell growth, and the testing method.

Abstract: Methods, compositions and materials useful in the detection of organophosphorous and organosulfur compounds are disclosed. In particular, biosensors wherein a porous or a non-porous support having an enzyme immobilized upon or within are disclosed. The biosensors exhibit enzymatic stability at extreme temperatures and/or denaturing conditions, and similar kinetic characteristics of the soluble form of the enzymes utilized. The enzyme does not leach from the porous or non-porous support and the material retains enzymatic activity after prolonged storage. Differential biosensors are also disclosed.

Abstract: The present invention provides a combination of reagent compositions for measuring an electrolyte which are excellent in stability, precision and quantitativity and have high solution stability sufficient to withstand distribution. In the combination of the reagent compositions of the present invention, a chelating agent and an inactivated &agr;-amylase capable of being reversibly activated by the electrolyte are formulated separately from each other.

Abstract: This invention provides modified phycobilisomes and phycobilisome complexes that are supramolecular complexes with diverse spectral properties, and which may optionally be immobilized on a manufactured solid support. The invention provides a versatile set of highly sensitive signal-generating systems and conjugates that may be used as highly detectable tracers and labels, or in biotransducers comprising phycobilisomes or phycobilisome complexes, and also provides methods for performing specific binding assays using signal-generating systems comprising phycobilisomes as detectable labels. The embodiments of the invention provide the art with an extremely sensitive, nonisotopic detection means for assaying analytes and for sensing molecular events and environmental conditions.

Abstract: Methods of measuring carboxypeptidase A levels and total carboxypeptidase A levels, wherein procarboxypeptidase A is converted to carboxypeptidase A by addition of clostripain, in a biological fluid with a carboxypeptidase A substrate, specificity of which is enhanced by addition of a carboxypeptidase A specific inhibitor are provided. In addition, methods of diagnosing acute pancreatitis by measurement of carboxypeptidase A levels and pancreatic cancer by measurement of total carboxypeptidase A levels are also provided.

Abstract: A method for measuring an enzyme reaction to determine an amount of a substance involved in the enzyme reaction, which comprises measuring a time course of a parameter of the enzyme reaction, measuring a time required for the parameter of the enzyme reaction to change from a first threshold value to a second threshold value, and correlating the measured time to an amount of the substance involved in the enzyme reaction.

Abstract: An improved homogeneous enzyme immunoassay process for quantitatively analyzing an antigen by determining the change in the enzymatic activity caused by a reaction between the antigen and an enzyme-labeled antibody. The antigen is reacted with an enzyme-labeled antibody, followed by the reaction with a second antibody capable of recognizing and binding to a different epitope and then with a third antibody capable of recognizing and binding to the second antibody. The enzymatic activity of the labeling enzyme is determined by a water-insoluble substrate. Using the water-insoluble substrate, steric hindrance is enhanced. A highly-sensitive analysis can be carried out by a simple operation even when the antigen has a molecular weight falling within an intermediate range, for example, a range of M.W. 10,000 to 70,000.

Abstract: A method of improving a phenotypic defect in a cell that contains a conformationally defective target protein wherein the conformational defect causes the phenotype defect, comprising contacting a first cell that expresses said confonnationally defective target protein with an amount of a protein stabilizing agent that is effective to improve the conformational defect, thereby improving the phenotypic defect of the first cell in comparison with a second cell having the same conformationally defective target protein and phenotypic defect, wherein the second cell is not contacted with the protein stabilizing agent.

Abstract: An aqueous protein solution buffered with a potassium phosphate buffer, in which the ratio of potassium ions to sodium ions in the solution is at least 10:1, is resistant to the formation of protein aggregates and particles under conditions of freezing, thawing, lyophilization, and reconstitution.

Abstract: The present invention relates to an enzyme sensor for measuring the concentration or activity of an analyte in a test fluid. The sensor has at least one enzyme layer comprising an immobilized enzyme for which the analyte is a substrate. The immobilized enzyme is obtained by formation of one or more covalent link(s), optionally by using a cross-linking agent, between the enzyme and at least one type of macromolecule in the presence of a competitive inhibitor for said enzyme. The present invention also relates to a membrane for an enzyme sensor. Furthermore, the invention relates to a method for stabilizing the enzymatic activity of an enzyme sensor.

Abstract: The invention provides sets of liquid reagents that afford calibration stability in an enzyme based spectrophotometric assay for the measurement of lactate in patient samples. The reagent sets include a lactate oxidase, a peroxidase, a hydrogen donor, an agent that substantially prevents ascorbic acid interference, and agent that substantially prevents bilirubin interference, a coupling agent, a buffer and, optionally, a preservative. The invention further provides methods for using the liquid reagent sets.

Abstract: The invention concerns a method of stabilizing molecules, or parts of molecules, which are sensitive to hydrolysis, in particular hydrolysis-sensitive labels or label-containing tracers in aqueous solutions. The invention also concerns kits for carrying out immunological assays making use of the method proposed.

Abstract: This invention relates to a method for evaluating the susceptibility of pharmaceutical drugs to metabolism by a specific cytochrome P-450 isozyme, which comprises contacting the sample compound with a reagent composition prepared by adding the specific cytochrome P-450 isozyme to liver microsomes lacking said specific cytochrome P-450 isozyme in a carrier material. More particularly, this invention relates to a useful in vitro quantitative assay for drug metabolism by P-450 isozymes such as CYP2D6, CYP2C19, and CYP2A6. This invention also relates to a reagent composition which is useful for said in vitro assay.

Abstract: The method of the invention involves providing a first receptacle and a second receptacle. The first receptacle contains a sterile aqueous broth and the second receptacle contains an aqueous broth including a carbon source. The method then includes placing into the first receptacle a first support surface having a paraffin wax coating thereon and placing into the second receptacle a second support surface having a hydrophobic material coating thereon. A body specimen, such as sputum, is then introduced into each of the first and second receptacles. The presence of a nonparaffinophilic hydrophobic microorganism in the body specimen is determined by observing (i) a lack of microorganism growth on the paraffin coated material of the first support surface and (ii) a presence of microorganism growth on the hydrophobic material coating of the second support surface. The presence of the nonparaffinophilic hydrophobic microorganism can be further confirmed by performing a DNA extraction.

Abstract: An assay device comprising a centrifugation tube and, as a sliding fit therein, a less deep inner tube whose base has one or more apertures sufficiently large to allow the passage of particles. This device can be used in an assay for microorganisms in a liquid sample containing fatty material, which comprises centrifuging the sample and a clearing agent in the device, removing the inner tube containing the fatty material, removing at least substantially all of the liquid supernatant in the centrifugation tube, and determining the presence of ATP in the sedimented pellet.

Abstract: Resistance of given plant tissue to an inhibitor of acetolactate synthase is identified by(a) combining in an aqueous medium a sample of the plant tissue, the inhibitor of acetolactate synthase, and an inhibitor of keto acid reductoisomerase;(b) allowing time for acetolactate to accumulate;(c) rupturing the cells;(d) acidifying the mixture to convert any accumulated acetolactate to acetoin; and(e) calorimetrically detecting the presence of acetoin in the mixture.

Abstract: A diagnostic method is provided for inflammatory bowel disorders (IBD), based on the relative levels of eosinophil granule proteins in physiological samples obtained from the GI tract of mammals suspected of having an IBD.

Abstract: The present invention relates to a dry immunoassay analytical element for assaying a ligand, comprising a support bearing: (a) a label zone comprising an enzyme labeled ligand or an enzyme labeled receptor; (b) a spreading zone; (c) a receptor zone comprising a fixed concentration of an immobilized receptor for the ligand; and (d) a gravure zone comprising a diaryl telluride. A prefered embodiment of the present invention further comprises a vanadyl salt. The present invention further relates to a method for performing an assay using an element as described above.

Abstract: A heat shock protein of Streptococcus pneumoniae named HSP72 and immunologically related polypeptides, the nucleotide and derived amino acid sequences of HSP72 (SEQ ID NO:4; SEQ ID NO:5), antibodies that bind to HSP72, and recombinant DNA methods for the production of HSP72 and immunologically related polypeptides. The polypeptides, DNA sequences and antibodies of this invention provide new means for the diagnosis, prevention and/or treatment of disease.

Abstract: This invention relates to an improved method for detecting and quantifying the presence of a target molecule, such as an antigen, an antibody or a polynucleotide, in a sample which method uses alkaline phosphatase as the reporter enzyme and the reduction of a tetrazolium salt to a formazan as part of the detection/signaling system.

Abstract: A device for performing an enzyme-labelled binding assay comprises an absorbent material and a developing solution containing a substrate for said enzyme, wherein the absorbent material is provided with a plurality of reagent zones including an indicator reagent zone and is capable of transporting the developing solution by capillary action sequentially through the reagent zones, and wherein the indicator reagent zone includes a reagent capable, directly or indirectly, of immobilizing an enzyme-labelled reagent in an amount dependent on the assay result, characterized in that the absorbent material includes a reagent that prevents a signal formation except where enzyme-labelled reagent is immobilized at the indicator reagent zone. The absorbent material is suitably in the form of an elongate strip provided with transverse reagent zones. The device is useful for performing immunoassays, including immunometric assays and dual analyte assays.

Abstract: This invention provides a method for measuring the concentration of urinary trypsin inhibitors which is excellent in precision and reproducibility, whose operation is simple, and in which a possibility of damaging a plastic cell is eliminated. The method for measuring the concentration of urinary trypsin inhibitors comprises mixing an urine sample, a protease solution containing trypsin, and a buffer solution, adding a substrate solution to the mixture to cause the enzyme reaction, and measuring the activity of the enzyme, wherein the buffer solution is prepared so that it contains at least 0.15 .mu.mol calcium per 1 .mu.g of the trypsin but no more than 100 .mu.

Abstract: An in vivo method for depleting mammalian cells of adenosine 5'-monophosphate (AMP) useful in the treatment of certain cancers is provided. According to the method, a population of cells is obtained from a host and assayed for loss of methylthioadenosine phosphorylase (MTAse) activity. MTAse catabolizes methylthioadenosine to adenine for endogenous salvage incorporation into the intracellular AMP pool. The preferred method for assaying loss of MTAse activity is a hybridization technique for detection of a homozygous loss of the gene which encodes MTAse. Hosts having MTAse deficient tumors are treated with a therapeutically effective amount of an agent which inhibits the activity of adenylsuccinate synthetase, which converts inosine 5'-monophosphate to AMP, thus depleting the tumor cells of substrates for de novo AMP production. L-alanosine is the preferred ASS inhibitory agent for use in the method of the invention.

Abstract: The present invention provides a method of improving the sensitivity and accuracy of a lead assay. The method enhances the recovery of lead during isolation of the lead from interfering compounds by maintaining the lead in a sample solution and making the recovered lead available for detection by the assay. An enhancing reagent complexes with the lead isolated in the sample solution. The enhancer includes a chelator having a lead equilibrium binding constant in the range of about 4 log K to about 13 log K. A kit for performing such a lead assay is also provided.

Abstract: This invention relates to a reagent for enzymatic determination of an analyte concentration in a patient wherein the degree of oxidation of a coenzyme is measured, characterized in that said reagent is stabilized against oxidation by a coenzyme reduction system comprising an enzyme and substrate pair selected so as to enable continuous regeneration of said coenzyme throughout storage of said reagent. Also disclosed is an improvement in an enzymatic method of determination of an analyte concentration in a sample body fluid wherein the degree of oxidation of a coenzyme is measured, the improvement comprising stabilizing a reagent comprising said coenzyme against oxidation by a coenzyme reduction system comprising an enzyme and substrate pair selected so as to enable continuous regeneration of said coenzyme throughout storage of said reagent. Also disclosed are reagents for the determination of aspartate aminotransferase, alanine aminotransferase, ammonia and urea.

Abstract: A bacteria impermeable container or ampule (10) contains a liquid growth medium and a substrate-indicator complex. The complex includes a substrate component, e.g., starch, and an indicator molecule, e.g., a dye, a fluorescent molecule, or the like, which are tightly bound and complexed, but which are cleavable by a preselected enzyme. A sterilant passes over a carrier (20) for microorganisms which, upon germination, are capable of rapidly generating large quantities of the preselected enzyme. Following the sterilization process, the carrier is immersed in the liquid growth medium. Any viable surviving microorganisms grow, generating the preselected enzyme. The enzymes cleave the bound indicator molecule from the substrate, resulting in a measurable property change in a couple of hours. Typical property changes include fluorescence, a color change, a change in pH which triggers a pH indicator color change, and the like.

Abstract: The invention relates to a composition comprising among others a fibrinopeptide A releasing compound. Furthermore the invention relates to the use of the composition as calibrator in plasma containing fibrinogen. A test kit comprising the said composition and a method to determine soluble fibrin also belong to the invention.

Abstract: Protein kinase mutant and wild-type genes encoding polypeptides of the class heretofore designated "casein kinase I" and useful in screening compositions which may affect DNA double-strand break repair activity are disclosed. Also disclosed are methods using the polynucleotides in cell-proliferative disorders.

Abstract: The present invention relates to single and dual-reporter luminescence assays utilizing general and specific reagents to quench enzyme-mediated reactions. In one embodiment of the invention, a reagent is added to the assay which non-specifically quenches enzyme-mediated luminescent reactions. In another embodiment of the invention, a reagent is added to the assay which simultaneously quenches one enzyme-mediated luminescent reaction while activating another distinct enzyme-mediated luminescent reaction. An assay kit containing specific quench reagents, and the reagents themselves are also disclosed.

Abstract: Disclosed herein is a cell containing a modified peptide. More specifically, the N-terminal amino acid residue of the peptide is modified by the addition of an aryl ketone group which, when contacted with an appropriate substrate, and exposed to light having a wavelength of about 330 nm or greater, results in the covalent bonding of the peptide to the substrate by a C--H insertion dominant mechanism. In preferred, embodiments, the aryl ketone is a benzophenone moiety. The peptide can be designed to specifically bind to a protein of interest in the cell. The cell is then contacted with light having a wavelength of greater than about 330 nm to bind the peptide covalently to the binding site on the intracellular protein of interest. In this way, the modified peptide can be used to specifically and irreversibly block a binding site on an intracellular protein of interest.

Abstract: Disclosed is an apparatus for processing body fluids which in one embodiment includes a support having ethylenediamine tetraacetic acid, and N-tosyl-lysyl chloromethylketone and/or N-tosyl-phenylalanyl chloromethylketone.

Abstract: An allergen immunoassay method features the use of a combination of a) closely controlled 1) elevated temperatures for assay reactions, 2) low temperatures for reagents and samples, 3) times for assay steps and especially assay reaction times, 4) reagent concentrations, and 5) reagent amounts; b) the use of a fast and accurate method of sample preparation that removes dust and contaminants; c) the stabilization of samples to avoid auto- and antibody degradation and unwanted effects of sample contaminants; and d) the formation of a colored product to determine the amount of a specific allergen. This combination provides an assay that can be completed in a few hours while retaining the precision, accuracy, sensitivity and response curve of previous methods requiring much longer periods of time.

Abstract: The invention provides a reagent for enzymatic determination of serum bicarbonate levels in a patient wherein the degree of oxidation of a coenzyme is measured and said reagent is stabilized against oxidation by a coenzyme reduction system comprising an enzyme and substrate pair selected to enable continuous regeneration of said coenzyme throughout storage of said reagent. The invention also provides an improvement in an enzymatic method of determination of the concentration of serum bicarbonate in a sample body fluid wherein the degree of oxidation of a coenzyme is measured, the improvement comprising stabilizing a reagent which comprises the coenzyme against oxidation by a coenzyme reduction system comprising an enzyme and substrate pair selected so as to enable continuous regeneration of the coenzyme throughout storage of the reagent.

Abstract: Methods and compositions are provided for use of modified or intact phycobilisomes as extremely potent labels in sensitive monitoring kits (e.g., for blood contamination), specific binding assays (e.g., visual, photometric and fluorometric immunoassays) and optoelectronic devices (e.g., biosensors, photoelectric transducers).

Abstract: A method is disclosed for stabilizing a conjugate of an enzyme and a member of a specific binding pair (enzyme conjugate). The method comprises the step of combining the enzyme conjugate with an effective amount of an antibody for the enzyme where the antibody does not substantially inhibit the activity of the enzyme. The invention has application to assays for the determination of an analyte wherein enzyme conjugates are employed. The improvement comprises employing as a reagent in the assay an immune complex of an enzyme conjugate and an antibody for the enzyme where the antibody does not substantially inhibit the activity of the enzyme. Compositions comprising such an immune complex and kits comprising such an immune complex in packaged combination with other assay reagents are also disclosed.

Abstract: A dry immunoassay analytical element for assaying a ligand, comprising a support bearing:1. an enzyme labeled ligand or an enzyme labeled receptor zone;2. a spreading zone; and3. a receptor zone containing a fixed concentration of an immobilized receptor for the ligand and the labeled ligand when present and the receptor is covalently bonded to polymeric beads having a diameter in the range of 0.1 to 5 .mu.m; characterized in that the element contains a diaryl telluride (DAT) compound and the zones can be in the same or separate layers.

Abstract: A method for increasing the binding activity of specific binding members bound to a solid phase material, e.g., a particle, that has been sterically stabilized. This increase in binding activity is brought about by degrading a steric stabilizer on the surface of the solid phase material. The method involves both immobilizing a specific binding member on the surface of a solid phase material and degrading a steric stabilizer on the surface of that solid phase material. In the preferred embodiment, the method involves the immobilization of a specific binding member on the surface of the sterically stabilized solid phase material, with subsequent degradation of the steric stabilizer.