Abstract: The invention provides methods and compositions for simultaneously detecting the activation state of a plurality of proteins in single cells using flow cytometry. The invention further provides methods and compositions of screening for bioactive agents capable of coordinately modulating the activity of a plurality of proteins in single cells. The methods and compositions can be used to determine the protein activation profile of a cell for predicting or diagnosing a disease state, and for monitoring treatment of a disease state.

Abstract: Bodily fluid is analyzed for the presence of drugs of a selected panel of drugs in a simultaneous assay in which sample of the fluid is incubated with additional amounts of all drugs of the panel, antibodies specific to each of the drugs of the panel, and microparticles, the microparticles being divided into subsets, one subset for each drug in the panel and each subset distinguishable from the others. The incubation is performed in a liquid medium in which competitive binding occurs, the drugs in the sample competing with those added to the assay medium for binding to the antibodies. In one procedure, the added drugs are pre-coupled to the microparticles while the antibodies are not, and the incubation is followed by further incubating the microparticles with labeled ligands that have affinity for the antibodies.

Abstract: Assay systems and specialized antibodies for the detection and quantitation of troponin I and troponin T in body fluids as an indicator of myocardial infarction are disclosed. Troponin I and T exist in various conformations and the ratios of monomeric troponin I and T and the binary and ternary complexes, as well as which form of troponin present in the blood, may be related to the metabolic state of the heart. Disclosed are systems to determine the presence of a troponin form or a group of troponin forms in whole blood, serum or plasma samples. Disclosed is a method for improving the recovery of troponin I or T from a surface used in immunoassays. Also disclosed are antibodies which recognize unbound troponin forms, the forms of troponin in binary complexes, the ternary complex of troponin I, T and C, and the conformations of troponin I having intramolecularly oxidized and reduced cysteines.

Abstract: The invention relates to Factor H gene polymorphisms and haplotypes associated with an elevated or a reduced risk of AMD. The invention provides methods and reagents for diagnosis and treatment of AMD.

Abstract: A method for the enumeration of micronucleated erythrocyte populations while distinguishing platelet and platelet-associated aggregates involves the use of a first fluorescent labeled antibody having binding specificity for a surface marker for reticulocytes, a second fluorescent labeled antibody having binding specificity for a surface marker for platelets, and a nucleic acid staining dye that stains DNA (micronuclei) in erythrocyte populations. Because the fluorescent emission spectra of the first and second fluorescent labeled antibodies do not substantially overlap with one another or with the emission spectra of the nucleic acid staining dye, upon excitation of the labels and dye it is possible to detect the fluorescent emission and light scatter produced by the erythrocyte populations and platelets, and count the number of cells from one or more erythrocyte populations in said sample.

Abstract: The invention is in the field of analytical technique and relates to an improved procedure for determining the concentration or activity of an analyte, an analyte-dependent detection reaction being set in motion and the reaction kinetics determined being evaluated. The procedure makes possible an individual determination of the lag phase of the reaction kinetics for each measurement.

Abstract: The current invention reports a method for producing an antibody comprising the steps of a) providing a plurality of hybridoma cells each expressing an antibody, b) determining the time dependent amount of said antibody bound to the respective antigen by surface plasmon resonance at different temperatures and different antibody concentrations, c) calculating with the time dependent amount determined in b) based on equations (II) to (XIII) at least the thermodynamic parameters (i) standard association binding entropy formula (A), (ii) standard dissociation binding entropy formula (B), (iii) standard binding entropy (?S°), (iv) free standard binding enthalpy (?G°), (v) standard dissociation free binding enthalpy formula (C), (vi) standard association free binding enthalpy formula (D), (vii) ?T?S°, (viii) dissociation rate constant kd, (ix) equilibrium binding constant KD, and (x) association rate constant ka, d) selecting a hybridoma cell producing an antibody with at least two of the following: i) a standard a

Abstract: A method of assaying nucleotide receptor P2X7 pore activity in white blood cells contained within a mixed cell sample is provided comprising labeling white blood cells with a white blood cell-specific label; depolarizing the labeled white blood cells with an isotonic depolarizing solution; contacting the labeled white blood cells with dye and a P2X7 agonist in an amount sufficient to activate P2X7 pore activity; contacting the labeled white blood cells with a divalent cation in an amount sufficient to deactivate P2X7 pore activity; and analyzing dye uptake whereby P2X7 pore activity is quantified by the amount of dye taken up in labeled white blood cells treated with the P2X7 agonist relative to labeled white blood cells in the absence of said P2X7 agonist, said P2X7 pore activity being corrected for sample age and by subtraction of P2X7 pore activity contributed by nonviable white blood cells.

Abstract: The present invention provides for methods and materials for diagnosing and treating neuromyelitis optica (NMO).

Abstract: Bodily fluid is analyzed for the presence of drugs of a selected panel of drugs in a simultaneous assay in which sample of the fluid is incubated with additional amounts of all drugs of the panel, antibodies specific to each of the drugs of the panel, and microparticles, the microparticles being divided into subsets, one subset for each drug in the panel and each subset distinguishable from the others. The incubation is performed in a liquid medium in which competitive binding occurs, the drugs in the sample competing with those added to the assay medium for binding to the antibodies. In one procedure, the added drugs are pre-coupled to the microparticles while the antibodies are not, and the incubation is followed by further incubating the microparticles with labeled ligands that have affinity for the antibodies.

Abstract: Whole cell, simultaneous target and drug-target assay using differentially labeled antibodies and flow cytometry. First antibody binds to total target and second antibody binds to the drug binding site of the target, thus drug binding will competitively inhibit the second antibody allowing for a competitive inhibition assay of drug-target binding. The assay allows for whole cell analysis and even analysis of mixed populations of cells, yet provides detailed kinetic assessment of drug activity.

Abstract: A method to determine specificity of ligand binding includes comparing a solid phase carrier first extract obtained by pre-treating a sample with a ligand-immobilized solid phase carrier and a solid phase carrier second extract obtained by treating the pretreated sample again with a ligand-immobilized solid phase carrier in terms of the proteins contained therein, and identifying a protein whose content is remarkably decreased in the second extract compared to the first extract, in order to solve 1) the problem of the solubility of subject ligand, 2) the problem of the non-specific protein-denaturing effect of the subject ligand added, and the like, in antagonism experiments in target search using an affinity resin.

Abstract: A method of detecting platelet thrombosis or organ failure in a patient suffering from disseminated intravascular coagulation (DIC) or systemic inflammatory response syndrome (SIRS), comprising analyzing a von Willebrand factor-cleaving protease and/or a cleaving factor thereof, is disclosed. A kit for detecting platelet thrombosis or organ failure in a patient suffering from DIC or SIRS, comprising an antibody or a fragment thereof which specifically binds to a von Willebrand factor-cleaving protease, and/or an antibody or a fragment thereof which specifically binds to a cleaving factor of the von Willebrand factor-cleaving protease, is disclosed.

Abstract: The present invention provides methods, diagnostic assays, and diagnostic kits based on said methods, to determine levels of immunosuppressive complexes containing immunosuppressive drugs tacrolimus, sirolimus and cyclosporine A separately and in combination, formed in the blood of a drug-treated patient or in a patient candidate to immunosuppressive drug therapy. These methods, assays and kits are especially useful when using automated systems.

Abstract: Methods and reagents for determining antithrombin III (AT) in body fluids by adding an AT binding partner to the sample and determining the free AT binding partner.

Abstract: According to the biosensor and the blood component analytical method of the present invention, in a biosensor that is made of a single layer or plural layers of a porous material as shown in FIG. 1, having a reagent holding part and utilizing chromatography, a cell shrinkage reagent is carried on at least part of the reagent holding part, or at least part of a chromatographically developed part that is upstream of the reagent holding part. According to the biosensor having the above-mentioned structure and the blood component analytical method, even when whole blood is a sample, a high-accuracy blood component analysis cart be performed easily and quickly with less cost.

Abstract: The invention concerns a free solution capillary electrophoresis method at alkaline pH for the analysis of samples comprising protein constituents including a lipoprotein constituent or constituents, characterized in that it comprises at least one step in which the sample is introduced into a capillary tube containing an analysis buffer, said analysis buffer further comprising at least one anionic surfactant type additive that is capable of hydrophobic interaction with the lipoprotein constituent(s) and of modifying the electrophoretic mobility. The invention also concerns a composition for capillary electrophoresis and a kit for analyzing protein constituents.

Abstract: The invention relates to an analytical measuring and evaluation method for determining the interaction parameters between an analyte and a ligand, preferably in a biosensor. According to the inventive method, the concentration of the analyte is gradually changed at defined intervals ti and the initial association or dissociation rates or association and dissociation rate constants are determined. The invention further relates to a device for carrying out the inventive method.

Abstract: Analytes using an active assay may be detected by introducing an analyte solution containing a plurality of analytes to a lacquered membrane. The lacquered membrane may be a membrane having at least one surface treated with a layer of polymers. The lacquered membrane may be semi-permeable to nonanalytes. The layer of polymers may include cross-linked polymers. A plurality of probe molecules may be arrayed and immobilized on the lacquered membrane. An external force may be applied to the analyte solution to move the analytes towards the lacquered membrane. Movement may cause some or all of the analytes to bind to the lacquered membrane. In cases where probe molecules are presented, some or all of the analytes may bind to probe molecules. The direction of the external force may be reversed to remove unbound or weakly bound analytes. Bound analytes may be detected using known detection types.

Abstract: Photonic crystal (PC) sensors, and sensor arrays and sensing systems incorporating PC sensors are described which have integrated fluid containment and/or fluid handling structures. The PC sensors are further integrated into a sample handling device such as a microwell plate. Sensors and sensing systems of the present disclosure are capable of high throughput sensing of analytes in fluid samples, bulk refractive index detection, and label-free detection of a range of molecules, including biomolecules and therapeutic candidates. The present disclosure also provides a commercially attractive fabrication platform for making photonic crystal sensors and systems wherein an integrated fluid containment structure and a photonic crystal structure are fabricated in a single molding or imprinting processing step amendable to high throughput processing.

Abstract: Disclosed are methods for conducting assays of samples, such as whole blood, that may contain cells or other particulate matter. Also disclosed are systems, devices, equipment, kits and reagents for use in such methods. One advantage of certain disclosed methods and systems is the ability to rapidly measure the concentration of an analyte of interest in blood plasma from a whole blood sample without blood separation and hematocrit correction.

Abstract: Heart and brain fatty acid binding proteins (H-FABP, B-FABP) are markers for stroke. The invention provides a diagnostic assay for either of these markers, preferably by ELISA using a anti-H-FABP or B-FABP antibody. Since H-FABP is also a marker for acute myocardial infarction (AMI), to distinguish stroke from AMI requires an assay specific to AMI, e.g. using troponin-1 or CK-MB as a marker, also to be carried out.

Abstract: A cell isolating device and method is provided to concentrate or isolate cells with specific characteristics from a mixture of different cell types. One embodiment may comprise two subtypes of antibodies that are directly conjugated to biotin (Abb) and conjugated to a fluorescent molecule (Abf). The conjugated antibodies (Abb+Abf) bind to the target cells in a mixed cell suspension. The cell suspension is then passed over an immobilized avidin or streptavidin substrate on a glass microscope slide. The biotinylated target cells adhere to the avidin/streptavidin substrate, while the unbound cells are washed off and collected in a wicking member. Captured cells on the avidin/streptavidin substrate may then be visualized directly using a fluorescent microscope or detected and enumerated via an on-board fluorescent detection device.

Abstract: The present invention provides methods and systems for performing in vivo flow cytometry. In one embodiments, selected circulating cells of interest of a subject are labeled with fluorescent probe molecules. The labeled cells are irradiated in-vivo so as to excite the fluorescent probes, and the radiation emitted by the excited probes is detected, preferably confocally. The detected radiation is then analyzed to derive desired information, such as relative cell count, of the cells of interest. In some embodiments, the circulating cells comprise apoptotic cells whose detection can allow, e.g., non-invasive monitoring of the efficacy of a cancer treatment, such as an anti-tumor or an anti-angiogenic therapy.

Abstract: Disclosed is a biomolecule chip and a fabrication method thereof. The biomolecule chip of the invention includes: a substrate; an insulating layer formed on the substrate; an adhesive layer formed on the insulating layer; a seed layer formed on the adhesive layer; an opening patterned at a predetermined location within the adhesive layer, the seed layer and the electroplating layer; and a biomolecule immobilized layer formed on the electroplating layer, the electroplating layer comprising a plasma-treated electroplating layer prior to the formation of the biomolecule immobilized layer. Accordingly, the immobilization of biomolecules onto the surface can be done more effectively by modifying the surface of the substrate in favor of biomolecules.

Abstract: This invention describes a method of using controlled fluidic forces to improve the performance of a biochemical binding assay where a target molecule is captured by specific molecular recognition onto a substrate surface with an affinity coating, and then labeled with a detectable micrometer-scale particle using a second specific molecular recognition reaction with the target. By using specific ranges of label sizes and laminar flow conditions, controlled fluidic forces can be applied to the label particles in order to selectively remove molecules bound to a surface according to their binding strength, and thereby increase the ratio of specifically bound labels to more weakly attached non-specifically bound labels. This method can be used with a wide variety of label types and associated detection methods, improving the sensitivity and selectivity of a broad range of binding assays.

Abstract: The present invention provides a method of performing a competitive assay for the detection and quantification of an analyte over an extended dynamic range. This is achieved by a multi-step sample addition method whereby different concentrations of sample are added at different times during the assay that produces a dose-response curve with multiple windows of detection. This multi-step sample addition method causes the dose-response curve of the composite assay to broaden, dramatically increasing the assay dynamic range.

Abstract: The invention provides methods and compositions for simultaneously detecting the activation state of a plurality of proteins in single cells using flow cytometry. The invention further provides methods and compositions of screening for bioactive agents capable of coordinately modulating the activity of a plurality of proteins in single cells. The methods and compositions can be used to determine the protein activation profile of a cell for predicting or diagnosing a disease state, and for monitoring treatment of a disease state.

Abstract: Disclosed are high throughput assay systems and methods for identifying agents that alter the level of expression of proteins in mammalian cells, particularly integral membrane proteins.

Abstract: Test strips designed to prevent or reduce false results when measuring the condition of a sample are provided. The test strips can be used for analysis of samples by methods including electrical and optical measurements. The reagent test strips and methods are particularly suited for use in the detection of blood coagulation.

Abstract: Damage to tissue, such as ischemic damage, can cause the release of mitochondrial proteins. The released proteins can be detected in a sample taken from a subject, indicating that the subject has suffered damage.

Abstract: The present invention provides methods of predicting or detecting labor in a female subject and methods of testing a compound for an ability to delay the onset of labor. The present invention also provides methods of testing a labor marker useful in the diagnostic methods, isolated peptides identified in the present invention, methods for inhibiting labor, utilizing the peptides, and kits comprising methods of the present invention.

Abstract: A method of and kit for diagnosing cardiovascular disease in a human involves assessing the presence and/or concentration of antibodies to platelet activating factor (PAF) in a sample of body fluid of the human.

Abstract: An assay technique for label-free, highly parallel, qualitative and quantitative detection of specific cell populations in a sample and for assessing cell functional status, cell-cell interactions and cellular responses to drugs, environmental toxins, bacteria, viruses and other factors that may affect cell function.

Abstract: There is provided a method of forming a multimeric complex having affinity for a target. The method comprises: obtaining a plurality of self-assembly molecules, said self-assembly molecules including complementary self-assembly units such as verotoxin subunit B, each of which is operatively connected to an interaction domain such as a single domain antibody specific for the target; and combining said self-assembly molecules such that at least three said self-assembly units simultaneously bind to one another so as to permit the single domain antibodies to bind the target.

Abstract: Novel, sensitive and specific markers for diagnostics and monitoring of liver injuries, including, but not limited to ischemic liver damage, are provided. This includes identification several enzymes of arginine/urea/nitric oxide cycle, sulfuration enzymes and spectrin breakdown related products, among others.

Abstract: A method of lowering the rate of a specific binding reaction in an assay for the detection and/or measurement of an analyte of interest is provided herein. In particular, the method includes providing a fluorescent conjugate of the analyte; a component capable of specifically binding to the analyte and its fluorescent conjugate; and a sample, which includes or is suspected to include the analyte. The method also includes allowing the specific binding component to interact simultaneously or at different times with the fluorescent conjugate of the analyte and the analyte in the sample, thereby forming a detectable complex due to the reaction between the fluorescent conjugate of the analyte and its specific binding component, wherein the reaction is performed in the presence of non-physiological amounts of at least one additive. The method further includes monitoring for the rate of change of the concentration of the detectable complex as a function of the amount of analyte in the sample.

Abstract: The present invention relates to a method for detecting von-Willebrand factor (vWF) activity comprising assaying a sample in the presence of a soluble form or portion of glycoprotein Ib(?) (GPIb(?) and ristocetin, or a functionally equivalent substance. Additionally, the invention relates to the use of the aforementioned soluble form or portion of glycoprotein Ib(?), of ristocetin or a functional equivalent substance, of specifically reacting anti-GPIB(?) antibody(ies) and/or of specific binding partners, like specifically reacting anti-vWF antibody(ies) for carrying out the method of the invention. Furthermore, the present invention relates to kits for carrying out the method of the invention.

Abstract: Disclosed are methods for detecting antibody in a sample, where the antibody targets an antigen expressed by red blood cells or red blood cell ghosts. Rather than detecting the binding events between a particular antigen antibody pair (as in traditional agglutination based assays) the methods herein allow for multiplexed detection of clinically important allo-immune antibodies to blood group antigens. Specifically the method involves generating fluorescently encoded red blood cells or red blood cell ghosts with known antigen presentation and using them to detect the presence of antibody in serum/plasma with a fluorescent sandwich type immunoassay. The assay results can be read using flow cytometric or fluorescent microscope based imaging techniques.

Abstract: In a process for the quantitative optical analysis of fluorescently labelled biological cells 5, a cell layer on a transparent support at the bottom 2 of a reaction vessel 1 is in contact with a solution 3 containing the fluorescent dye 4. The sensitivity of analytical detection can be considerably improved if to the fluorescent dye 4 already present in addition a masking dye 9, which absorbs the excitation light 6 for the fluorescent dye 4 and/or its emission light 7, is added to the solution 3 and/or if a separating layer 10 permeable to the solution and absorbing and/or reflecting the excitation light 6 or the emission light 7 is applied to the cell layer at the bottom 2. This process can also be used for improving the sensitivity in the quantitative optical analysis of a luminescent biological cell layer. The separating layer 10 must in this case be composed such that it has a high power of reflection for the luminescent light 11.

Abstract: The present invention relates to fluorescence polarization assays for determining the HDC modulating activity of a candidate compound including providing a reaction mixture comprising a HDC, histidine, a fluorescently labeled histamine probe, a candidate compound and an anti histamine antibody having selectivity for histamine at least 10 fold greater than histidine: incubating the reaction mixture; and determining whether inhibition of HDC has occurred in the presence of the test compound, wherein an increase in fluorescence signal is an indication that the test compound inhibits the activity of the HDC.

Abstract: A method having clinically sufficient degree of diagnostic accuracy for detecting the presence of coronary artery disease in a human patient from the general population and for distinguishing between the stages of the disease in that patient is disclosed. The stages are, first, the non-acute stage, which is either asymptomatic coronary artery disease or stable angina, second, the acute stage known as unstable angina, and, third, the acute stage known as acute myocardial infarction. The diseased state (as opposed to the non-diseased state) is indicated by the clinically significant presence of a first marker in a sample from the patient. The presence of one of the two acute stages, unstable angina or acute myocardial infarction, is indicated by the clinically significant presence of a second marker in a sample from the patient. The presence of the more severe acute stage known as acute myocardial infarction is indicated by the clinically significant presence of a third marker in a sample from the patient.

Abstract: Assay systems and specialized antibodies for the detection and quantitation of troponin I and troponin T in body fluids as an indicator of myocardial infarction. Since troponin I and T exist in various conformations in the blood, the ratios of the monomeric troponin I an T and the binary and ternary complexes, as well as which form of troponin present in the blood, may be related to the metabolic state of the heart. Disclosed is a system to determine the presence of a troponin form or a group of troponin forms in a sample of whole blood, serum or plasma. Disclosed is a stabilized composition of troponin; the stabilized composition can comprise a stabilized composition of troponin I, wherein the troponin I is oxidized, the troponin I can be unbound or the troponin I can be in a complex.

Abstract: The present invention provides methods, diagnostic assays, and diagnostic kits based on said methods, to determine levels of immunosuppressive complexes containing immunosuppressive drugs tacrolimus, sirolimus and cyclosporine A separately and in combination, formed in the blood of a drug-treated patient or in a patient candidate to immunosuppressive drug therapy. These methods, assays and kits are especially useful when using automated systems.

Abstract: A method to determine specificity of ligand binding includes comparing a solid phase carrier first extract obtained by pre-treating a sample with a ligand-immobilized solid phase carrier and a solid phase carrier second extract obtained by treating the pretreated sample again with a ligand-immobilized solid phase carrier in terms of the proteins contained therein, and identifying a protein whose content is remarkably decreased in the second extract compared to the first extract, in order to solve 1) the problem of the solubility of subject ligand, 2) the problem of the non-specific protein-denaturing effect of the subject ligand added, and the like, in antagonism experiments in target search using an affinity resin.

Abstract: A method of measuring the rate of binding of a binding substance and an analyte, for example in an assay such as an immunoassay, uses an initial step of performing ultrasonication sufficient to disrupt binding between the binding substance and the analyte. After cessation of the ultrasonication, measurements are taken to determine the rate of binding at cessation of said ultrasonication or at a predetermined time thereafter The ultrasonication results in knowledge of the precise time of the start of the binding reaction which provides a better rate measurement.

Abstract: Heterogeneous assays for different analytes in a single biological sample are performed simultaneously in a multiplexed assay that combines flow cytometry with the use of magnetic particles as the solid phase and yields an individual result for each analyte. The particles are distinguishable from each other by characteristics that permit them to be differentiated into groups, each group carrying an assay reagent bonded to the particle surface that is distinct from the assay reagents of particles in other groups. The magnetic particles facilitate separation of the solid and liquid phases, permitting the assays to be performed by automated equipment. Assays are also disclosed for the simultaneous detection of antibodies of different classes and a common antigen specificity or of a common class and different antigen specificities.

Abstract: A general immunoglobulin-target assay system is provided, in which a positive outcome (the generation of a signal) depends only on the intracellular interaction of immunoglobulin with target. This can be accomplished for many immunoglobulins expressed in yeast and/or in mammalian cells and allows the selection of immunoglobulins which are capable of functioning in an intracellular environment.

Abstract: The invention provides methods and compositions for simultaneously detecting the activation state of a plurality of proteins in single cells using flow cytometry. The invention further provides methods and compositions of screening for bioactive agents capable of coordinately modulating the activity of a plurality of proteins in single cells. The methods and compositions can be used to determine the protein activation profile of a cell for predicting or diagnosing a disease state, and for monitoring treatment of a disease state.

Abstract: The present invention relates to a method for analysing a cell sample for cell surface-bound or intracellularly bound analytes by providing an array of immobilised specific binding agents for a set of different ligands, where each ligand is specific to a respective cell surface-bound or intracellularly bound analyte, and using the array and the set of ligands in an inhibition type or a direct type assay format to determine cell surface-bound or intracellularly bound analytes in the cell sample. The invention also relates to assay kits for cell characterization.