Syva Company

International Diagnostic Technology

Edward T Maggio

Edwin F Ullman

Robert F Zuk

Robert A Yoshida

Richard L Wife

Dietrich Rehbinder

Janet H Chen

Double tagged immunoassay

A competitive protein binding method is provided for the determination of an analyte which is a member of an immunological pair consisting of ligand and receptor for the ligand. A chemiluminescent source is employed comprised of one or more individual members, one chemiluminescent source member being conjugated to one of the members of the immunological pair, so as to provide chemiluminescence adjacent to the site of conjugation. A quencher molecule is conjugated to a member of the immunological pair. When the members of the immunological pair bind, the quencher molecule is brought within quenching distance of the chemiluminescent source so as to inhibit the emission of light by the chemiluminescent source. The amount of analyte present in the assay medium affects the amount of binding between the members of the immunological pair which results in quenching of the chemiluminescence. By observing the light emitted from the assay medium, either from the chemiluminescent source or the quencher, the change in light emission in relation to the concentration of analyte present in the assay medium can be used to determine the amount of analyte present in the assay medium. By employing standards having known amounts of analyte, the amount of analyte in an unknown sample can be quantitatively determined. Reagent kits can be provided having predetermined amounts of the reagents, so as to substantially optimize the sensitivity of the assay.

Kit for carrying out chemically induced fluorescence immunoassay

Methods and reagent combinations are provided for competitive protein binding assays for determining a member of an immunological pair (ligand and receptor) whereby an enzyme-ligand conjugate is employed in combination with an enzyme inhibitor, conveniently an antibody to said enzyme. When ligand is the analyte, receptor for ligand is also included in the assay medium, while supplemental amounts of receptor need not be added when receptor is the analyte. The assay is carried out in an aqueous buffered medium, normally at constant temperature, by combining in the assay medium the unknown sample suspected of containing the analyte, enzyme-bound-ligand, ligand receptor (antiligand), enzyme inhibitor (antienzyme), and enzyme substrates, and the enzymatic activity in the assay medium determined. By comparing the observed enzymatic activity with an unknown to the enzymatic activity observed in an assay medium with a known amount of analyte, the amount of analyte can be quantitatively determined. Kits are provided having matched amounts of enzyme-bound-ligand, antienzyme and, when appropriate, antiligand for use in the subject assay.

Antienzyme homogeneous competitive binding assay

Method and compositions are provided for chemical analysis of an analyte which is a member of a specific binding pair of organic substances consisting of ligand and ligand receptor (antiligand). The method involves bringing together the following reagents with the analyte in an aqueous assay medium under mild conditions. The first reagent is a conjugate of a member of the specific binding pair with a chemical entity which provides a means for chemically changing the concentration of a compound which acts as a signal mediator. The second reagent is the signal mediator precursor. The third reagent is a conjugate of a member of the specific binding pair with a component of a signal producing system of which system the signal mediator is a member. The amount of signal which can be detected is affected by the local concentration of the signal mediator. By bringing the reagents together in the presence of analyte, where the signal mediator concentration changing means is brought together in a microenvironment with the conjugated signal producing system component, localized concentrations of the signal mediator can be created which differ from the gross concentration of the signal mediator in the assay medium. The degree to which the signal mediator concentration changing means is in close proximity to the signal producing means in a microenvironment will affect the observed signal. By appropriate choice of the two conjugates in conjunction with the analyte, the observed signal can be related to the amount of analyte in the medium. Novel conjugates are provided, as well as combinations of conjugates in specific proportions to substantially optimize the assay sensitivity. The combinations are provided as kits, where ancillary reagents can also be included, so as to simplify the combination of reagents, as well as provide for more accurate measurements and relative proportions of reagents.

Reagents and method employing channeling

A competitive protein binding method is provided for the determination of an analyte which is a member of an immunological pair consisting of ligand and receptor for the ligand. A chemiluminescent source is employed comprised of one or more individual members, one chemiluminescent source member being conjugated to one of the members of the immunological pair, so as to provide chemiluminescence adjacent to the site of conjugation. A quencher molecule is conjugated to a member of the immunological pair. When the members of the immunological pair bind, the quencher molecule is brought within quenching distance of the chemiluminescent source so as to inhibit the emission of light by the chemiluminescent source. The amount of analyte present in the assay medium affects the amount of binding between the members of the immunological pair which results in quenching of the chemiluminescence. By observing the light emitted from the assay medium, either from the chemiluminescent source of the quencher, the change in light emission in relation to the concentration of analyte present in the assay medium can be used to determine the amount of analyte present in the assay medium. By employing standards having known amounts of analyte, the amount of analyte in an unknown sample can be quantitatively determined. Reagent kits can be provided having predetermined amounts of the reagents, so as to substantially optimize the sensitivity of the assay.

Chemically induced fluorescence immunoassay

Methods and reagents are provided for immunoassays employing as reagents a labelled receptor, where the label is capable of providing a detectable signal, and modifying reagents, capable of modifying the signal obtained from the label. When ligand (ligand analog for monoepitopic ligands) is present in the assay medium, a complex is formed which inhibits interaction between the label modifying reagent and the label. By measuring the signal obtained in the presence of a known amount of ligand in the assay medium and comparing that signal with the signal obtained with an unknown sample suspected of containing ligand, one can qualitatively or quantitatively determine the amount of ligand in the unknown. For determinating anti(ligand) the assay is carried out in substantially the same way, except a source of ligand or ligand analog must be provided.

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Redwood City, CA, United States of America

Edward T Maggio