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: 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, 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: 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 said 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: 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 performing isothermal amplification of a nucleic acid target employing probes characterized by having a masked RNA polymer promoter unable to bind to a complementary initiator oligonucleotide and RNA polymerase and initiate transcription, a dsDNA sequence which when invaded by the target nucleic acid exposes the masked promoter to initiate transcription, and a template sequence, a portion of which is normally included in the dsDNA region, which when copied produces a product that can reinitiate the process of invading the dsDNA region and initiating transcription of another copy.