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Bioluminescent immunoassay using a monomeric Fab'-photoprotein aequorin conjugate. 1991

T Erikaku, and S Zenno, and S Inouye
Yokohama Research Center, Chisso Corporation, Japan.

The photoprotein aequorin emits light by an intermolecular reaction when mixed with Ca2+. To apply a bioluminescent immunoassay based on the light emission property of aequorin, we prepared aequorin-antibody Fab' conjugates by a chemical cross-linking technique. Recombinant apoaequorin was coupled with human tumor necrosis factor-alpha (TNF-alpha) antibodies (polyclonal Fab' or monoclonal Fab' fragments) using N-succinimidyl 4-(N-maleimidemethyl)cyclohexane-1-carboxylate. The luminescent activity of the Fab'-aequorin conjugate was about one-tenth that of aequorin. Using a monomeric conjugate, human TNF-alpha can be measured at an attomole level by a sandwich immunoassay technique.

UI MeSH Term Description Entries
D007118 Immunoassay A technique using antibodies for identifying or quantifying a substance. Usually the substance being studied serves as antigen both in antibody production and in measurement of antibody by the test substance. Immunochromatographic Assay,Assay, Immunochromatographic,Assays, Immunochromatographic,Immunoassays,Immunochromatographic Assays
D007140 Immunoglobulin Fab Fragments Univalent antigen-binding fragments composed of one entire IMMUNOGLOBULIN LIGHT CHAIN and the amino terminal end of one of the IMMUNOGLOBULIN HEAVY CHAINS from the hinge region, linked to each other by disulfide bonds. Fab contains the IMMUNOGLOBULIN VARIABLE REGIONS, which are part of the antigen-binding site, and the first IMMUNOGLOBULIN CONSTANT REGIONS. This fragment can be obtained by digestion of immunoglobulins with the proteolytic enzyme PAPAIN. Fab Fragment,Fab Fragments,Ig Fab Fragments,Immunoglobulins, Fab Fragment,Fab Immunoglobulin Fragments,Immunoglobulin Fab Fragment,Immunoglobulins, Fab,Fab Fragment Immunoglobulins,Fab Fragment, Immunoglobulin,Fab Fragments, Immunoglobulin,Fragment Immunoglobulins, Fab,Fragment, Fab,Immunoglobulin Fragments, Fab
D007202 Indicators and Reagents Substances used for the detection, identification, analysis, etc. of chemical, biological, or pathologic processes or conditions. Indicators are substances that change in physical appearance, e.g., color, at or approaching the endpoint of a chemical titration, e.g., on the passage between acidity and alkalinity. Reagents are substances used for the detection or determination of another substance by chemical or microscopical means, especially analysis. Types of reagents are precipitants, solvents, oxidizers, reducers, fluxes, and colorimetric reagents. (From Grant & Hackh's Chemical Dictionary, 5th ed, p301, p499) Indicator,Reagent,Reagents,Indicators,Reagents and Indicators
D008163 Luminescent Measurements Techniques used for determining the values of photometric parameters of light resulting from LUMINESCENCE. Bioluminescence Measurements,Bioluminescent Assays,Bioluminescent Measurements,Chemiluminescence Measurements,Chemiluminescent Assays,Chemiluminescent Measurements,Chemoluminescence Measurements,Luminescence Measurements,Luminescent Assays,Luminescent Techniques,Phosphorescence Measurements,Phosphorescent Assays,Phosphorescent Measurements,Assay, Bioluminescent,Assay, Chemiluminescent,Assay, Luminescent,Assay, Phosphorescent,Assays, Bioluminescent,Assays, Chemiluminescent,Assays, Luminescent,Assays, Phosphorescent,Bioluminescence Measurement,Bioluminescent Assay,Bioluminescent Measurement,Chemiluminescence Measurement,Chemiluminescent Assay,Chemiluminescent Measurement,Chemoluminescence Measurement,Luminescence Measurement,Luminescent Assay,Luminescent Measurement,Luminescent Technique,Measurement, Bioluminescence,Measurement, Bioluminescent,Measurement, Chemiluminescence,Measurement, Chemiluminescent,Measurement, Chemoluminescence,Measurement, Luminescence,Measurement, Luminescent,Measurement, Phosphorescence,Measurement, Phosphorescent,Measurements, Bioluminescence,Measurements, Bioluminescent,Measurements, Chemiluminescence,Measurements, Chemiluminescent,Measurements, Chemoluminescence,Measurements, Luminescence,Measurements, Luminescent,Measurements, Phosphorescence,Measurements, Phosphorescent,Phosphorescence Measurement,Phosphorescent Assay,Phosphorescent Measurement,Technique, Luminescent,Techniques, Luminescent
D008301 Maleimides Derivatives of maleimide (the structural formula H2C2(CO)2NH) containing a pyrroledione ring where the hydrogen atom of the NH group is replaced with aliphatic or aromatic groups.
D011994 Recombinant Proteins Proteins prepared by recombinant DNA technology. Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D002850 Chromatography, Gel Chromatography on non-ionic gels without regard to the mechanism of solute discrimination. Chromatography, Exclusion,Chromatography, Gel Permeation,Chromatography, Molecular Sieve,Gel Filtration,Gel Filtration Chromatography,Chromatography, Size Exclusion,Exclusion Chromatography,Gel Chromatography,Gel Permeation Chromatography,Molecular Sieve Chromatography,Chromatography, Gel Filtration,Exclusion Chromatography, Size,Filtration Chromatography, Gel,Filtration, Gel,Sieve Chromatography, Molecular,Size Exclusion Chromatography
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000331 Aequorin A photoprotein isolated from the bioluminescent jellyfish Aequorea. It emits visible light by an intramolecular reaction when a trace amount of calcium ion is added. The light-emitting moiety in the bioluminescence reaction is believed to be 2-amino-3-benzyl-5-(p-hydroxyphenyl)pyrazine (AF-350). Aequorine
D000911 Antibodies, Monoclonal Antibodies produced by a single clone of cells. Monoclonal Antibodies,Monoclonal Antibody,Antibody, Monoclonal

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