Abstract: Methods, compositions and kits are disclosed. The compositions are light emitting and comprise a polymeric matrix having dissolved therein a photoactive compound. The composition has the characteristic that, after activation of the photoactive compound, the rate of decrease in the intensity of light emission at any time during a 20-fold decrease in the intensity is proportional to the intensity of the light emission. In one embodiment the polymeric matrix is comprised of particles of about 20 nm to about 100 ?m in diameter to which is bound a specific binding pair member. The particles generally comprise a polymeric matrix having dissolved therein about 1 to about 20% by weight of a dopant. The compositions may be used in methods for determining an analyte. A combination is provided comprising (1) a medium suspected of containing the analyte, (2) and the aforementioned composition. The photoactive substance is activated and the effect of the activating on the optical properties of the combination is detected.

Abstract: Methods are disclosed for determining minute quantities of an analyte in a medium suspected of containing the analyte. One method comprises treating a medium suspected of containing an analyte under conditions such that the analyte, if present, causes a substrate having an oxidant cleavable linker and a photosensitizer to come into close proximity. The photosensitizer generates singlet oxygen which oxidatively cleaves the linker to form a product which can be detected in a sandwich detection assay such as LOCI. The amount of product detected is related to the amount of analyte in the medium. Compositions, kits, and compounds are also disclosed.

Abstract: A method is disclosed for modifying an oligonucleotide, which method has application to the detection of a polynucleotide analyte. An oligonucleotide is reversibly hybridized with a polynucleotide, for example, a polynucleotide analyte, in the presence of a 5?-nuclease under isothermal conditions. The polynucleotide analyte serves as a recognition element to enable a 5?-nuclease to cleave the oligonucleotide to provide (i) a first fragment that is substantially non-hybridizable to the polynucleotide analyte and (ii) a second fragment that lies 3? of the first fragment (in the intact oligonucleotide) and is substantially hybridizable to the polynucleotide analyte. At least a 100-fold molar excess of the first fragment and/or the second fragment are obtained relative to the molar amount of the polynucleotide analyte. The presence of the first fragment and/or the second fragment is detected, the presence thereof indicating the presence of the polynucleotide analyte.

Abstract: Methods and compositions are provided for determining ADP in the presence of ATP. These comprise including among the assay reagents at least one of the correcting components creatine phosphokinase and phosphocreatine, pyruvate kinase and phosphoenolpyruvate, peroxidase and a non-interfering peroxidase substrate, and catalase. One aspect of the method employs formation of hydrogen peroxide from the ADP by pyruvate kinase, phosphoenolpyruvate and pyruvate oxidase. The hydrogen peroxide is then determined. A combined reagent having all of the reagents may optionally include a peroxidase when the hydrogen peroxide is to be enzymatically determined. A peroxidase substrate is added to the sample in conjunction with the peroxidase substrate reagent, the mixture incubated and depending on whether the peroxidase substrate is a fluorescer or chemiluminescer, the mixture may be illuminated with excitation light and the emitted light determined as a measure of the ADP in the sample.

Abstract: A sensitive intracellular calcium assay is disclosed comprising conveniently a reagent comprised of a dye precursor capable of entering cells and being hydrolyzed to a dye, whereby the dye complexes with calcium in the cells and provides a luminescent signal, an antibody specific for the dye and conjugated with a quencher, and a cellular anion exchange enzyme inhibitor. In performing the assay, the reagent is combined with cells expressing a receptor responsive to a ligand resulting in a change in cytosolic calcium. After incubation for the dye precursor to permeate the cells, the calcium may be determined by exciting the dye precursor and determining the peak fluorescence over a time course. The method can be used for measuring the effect of an agent on cytosolic calcium by binding to a cell surface membrane receptor.

Abstract: Compositions are disclosed comprising (a) a metal chelate wherein the metal is selected from the group consisting of europium, terbium, dysprosium, samarium osmium and ruthenium in at least a hexacoordinated state and (b) a compound having a double bond substituted with two aryl groups, an oxygen atom and an atom selected from the group consisting of oxygen, sulfur and nitrogen wherein one of the aryl groups is electron donating with respect to the other. Such composition is preferably incorporated in a latex particulate material. Methods and kits are also disclosed for determining an analyte in a medium suspected of containing the analyte. The methods and kits employ as one component a composition as described above.

Abstract: Methods, compositions and kits are disclosed. The methods are directed to determining the presence of one or more analytes in a sample suspected of containing any one of a plurality of the analytes. A combination is provided comprising in a medium (i) the sample, (ii) a binding partner for each of the analytes, (iii) for each of the analytes, a first reagent comprising a member of a signal producing system, a ligand and an analyte analog, and (iv) a second reagent comprising a binding partner for the ligand. The binding of the binding partner for the ligand is affected by the presence of an analyte and alters the amount of signal produced by the member of the signal producing system. The signal thus is modulated if one or more of the analytes are present in the sample. The amount of the signal is determined and is related to the presence of one or more of the analytes in the sample. The method may be homogeneous or heterogeneous.

Abstract: Methods, compositions and kits are disclosed. The methods are directed to determining the presence of one or more analytes in a sample suspected of containing any one of a plurality of the analytes. A combination is provided comprising in a medium (i) the sample, (ii) a binding partner for each of the analytes, (iii) for each of the analytes, a first reagent comprising a member of a signal producing system, a ligand and an analyte analog, and (iv) a second reagent comprising a binding partner for the ligand. The binding of the binding partner for the ligand is affected by the presence of an analyte and alters the amount of signal produced by the member of the signal producing system. The signal thus is modulated if one or more of the analytes are present in the sample. The amount of the signal is determined and is related to the presence of one or more of the analytes in the sample. The method may be homogeneous or heterogeneous.

Abstract: Methods, compositions and kits are disclosed. The compositions are light emitting and comprise a polymeric matrix having dissolved therein a photoactive compound. The composition has the characteristic that, after activation of the photoactive compound, the rate of decrease in the intensity of light emission at any time during a 20-fold decrease in the intensity is proportional to the intensity of the light emission. In one embodiment the polymeric matrix is comprised of particles of about 20 nm to about 100 ?m in diameter to which is bound a specific binding pair member. The particles generally comprise a polymeric matrix having dissolved therein about 1 to about 20% by weight of a dopant. The compositions may be used in methods for determining an analyte. A combination is provided comprising (1) a medium suspected of containing the analyte, (2) and the aforementioned composition. The photoactive substance is activated and the effect of the activating on the optical properties of the combination is detected.

Abstract: Compositions are disclosed comprising (a) a metal chelate wherein the metal is selected from the group consisting of europium, terbium, dysprosium, samarium osmium and ruthenium in at least a hexacoordinated state and (b) a compound having a double bond substituted with two aryl groups, an oxygen atom and an atom selected from the group consisting of oxygen, sulfur and nitrogen wherein one of the aryl groups is electron donating with respect to the other. Such composition is preferably incorporated in a latex particulate material. Methods and kits are also disclosed for determining an analyte in a medium suspected of containing the analyte. The methods and kits employ as one component a composition as described above.

Abstract: Methods, compositions and kits are disclosed. The methods are directed to determining the presence of one or more analytes in a sample suspected of containing any one of a plurality of the analytes. A combination is provided comprising in a medium (i) the sample, (ii) a binding partner for each of the analytes, (iii) for each of the analytes, a first reagent comprising a member of a signal producing system, a ligand and an analyte analog, and (iv) a second reagent comprising a binding partner for the ligand. The binding of the binding partner for the ligand is affected by the presence of an analyte and alters the amount of signal produced by the member of the signal producing system. The signal thus is modulated if one or more of the analytes are present in the sample. The amount of the signal is determined and is related to the presence of one or more of the analytes in the sample. The method may be homogeneous or heterogeneous.

Abstract: Methods, compositions and kits are disclosed. The compositions are light emitting and comprise a polymeric matrix having dissolved therein a photoactive compound. The composition has the characteristic that, after activation of the photoactive compound, the rate of decrease in the intensity of light emission at any time during a 20-fold decrease in the intensity is proportional to the intensity of the light emission. In one embodiment the polymeric matrix is comprised of particles of about 20 nm to about 100 &mgr;m in diameter to which is bound a specific binding pair member. The particles generally comprise a polymeric matrix having dissolved therein about 1 to about 20% by weight of a dopant. The compositions may be used in methods for determining an analyte. A combination is provided comprising (1) a medium suspected of containing the analyte, (2) and the aforementioned composition.

Abstract: Alpha-haloketones are useful alkylating agents for coupling to sulfhydryl-containing biomolecules. However, they react spontaneously with water, alkali and organic bases and therefore cannot be stored for extended periods of time in aqueous solutions, particularly in the presence of proteins at physiological pH. The present invention provides novel solutions to these problems, however, as novel compounds and compositions comprising protected haloketones are disclosed herein. Methods of preparing and using protected haloketones which are useful in a variety of applications—e.g., in assays and conjugation reactions—are also disclosed herein.

Abstract: Methods, compositions and kits are disclosed. The methods are directed to determining the presence of one or more analytes in a sample suspected of containing any one of a plurality of the analytes. A combination is provided comprising in a medium (i) the sample, (ii) a binding partner for each of the analytes, (iii) for each of the analytes, a first reagent comprising a member of a signal producing system, a ligand and an analyte analog, and (iv) a second reagent comprising a binding partner for the ligand. The binding of the binding partner for the ligand is affected by the presence of an analyte and alters the amount of signal produced by the member of the signal producing system. The signal thus is modulated if one or more of the analytes are present in the sample. The amount of the signal is determined and is related to the presence of one or more of the analytes in the sample. The method may be homogeneous or heterogeneous.

Abstract: Methods are disclosed for determining minute quantities of an analyte in a medium suspected of containing the analyte. One method comprises treating a medium suspected of containing an analyte under conditions such that the analyte, if present, causes a substrate having an oxidant cleavable linker and a photosenstizer to come into close proximity. The photosenstizer generates singlet oxygen which oxidatively cleaves the linker to form a product which can be detected in a sandwich detection assay such as LOCI. The amount of product detected is related to the amount of analyte in the medium. Compositions, kits, and compounds are also disclosed.

Abstract: Alpha-haloketones are useful alkylating agents for coupling to sulfhydryl-containing biomolecules. However, they react spontaneously with water, alkali and organic bases and therefore cannot be stored for extended periods of time in aqueous solutions, particularly in the presence of proteins at physiological pH. The present invention provides novel solutions to these problems, however, as novel compounds and compositions comprising protected haloketones are disclosed herein. Methods of preparing and using protected haloketones which are useful in a variety of applications—e.g., in assays and conjugation reactions—are also disclosed herein.

Abstract: Compositions are disclosed comprising (a) a metal chelate wherein the metal is selected from the group consisting of europium, terbium, dysprosium, samarium osmium and ruthenium in at least a hexacoordinated state and (b) a compound having a double bond substituted with two aryl groups, an oxygen atom and an atom selected from the group consisting of oxygen, sulfur and nitrogen wherein one of the aryl groups is electron donating with respect to the other. Such composition is preferably incorporated in a latex particulate material. Methods and kits are also disclosed for determining an analyte in a medium suspected of containing the analyte. The methods and kits employ as one component a composition as described above.

Abstract: Methods, compositions and kits are disclosed. The compositions are light emitting and comprise a polymeric matrix having dissolved therein a photoactive compound. The composition has the characteristic that, after activation of the photoactive compound, the rate of decrease in the intensity of light emission at any time during a 20-fold decrease in the intensity is proportional to the intensity of the light emission. In one embodiment the polymeric matrix is comprised of particles of about 20 nm to about 100 &mgr;m in diameter to which is bound a specific binding pair member. The particles generally comprise a polymeric matrix having dissolved therein about 1 to about 20% by weight of a dopant. The compositions may be used in methods for determining an analyte. A combination is provided comprising (1) a medium suspected of containing the analyte, (2) and the aforementioned composition.

Abstract: Compositions are disclosed comprising (a) a metal chelate wherein the metal is selected from the group consisting of europium, terbium, dysprosium, samarium osmium and ruthenium in at least a hexacoordinated state and (b) a compound having a double bond substituted with two aryl groups, an oxygen atom and an atom selected from the group consisting of oxygen, sulfur and nitrogen wherein one of the aryl groups is electron donating with respect to the other. Such composition is preferably incorporated in a latex particulate material. Methods and kits are also disclosed for determining an analyte in a medium suspected of containing the analyte. The methods and kits employ as one component a composition as described above.

Abstract: Methods and compositions are provided for determining large numbers of single nucleotide polymorphisms in target DNA employing particles having (1) primers complementary to sequences in the target DNA where the next succeeding 3′-nucleotide is a potential single nucleotide polymorphism and coding composition members, where the members are unique for each primer, and (2) differentially labeled terminating nucleotides, where the label permits separation of the terminating nucleotides. Desirably the particles are separated into groups having a common prevalent next succeeding nucleotide. The particles and target DNA are combined under nucleotide extending conditions, the particles separated into groups in accordance with the terminating nucleotide and the coding members identified, so that one knows the sequence and the single nucleotide polymorphism. Various protocols are provided for the determination.