Photoaffinity Labeling of Antibodies for Applications in Homogeneous fluoroimmunoassays

A homogeneous noncompetitive immunoassay based on photoaffinity labeling techniques is described. Using this method, a fluorophore (reporter) can be specifically attached to an antibody in the vicinity of its antigen-combining sites. Upon antigen binding, changes in the fluorescence spectrum of the...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 1995-03, Vol.67 (5), p.959-966
Main Authors: Chang, I.-Nan, Lin, Jinn-Nan, Andrade, Joseph D, Herron, James N
Format: Article
Language:English
Subjects:
Affinity Labels
Antibodies - immunology
Biochemistry
Biological and medical sciences
Biotin - immunology
Dansyl Compounds - chemistry
Fluorescent Dyes - chemistry
Fluoroimmunoassay - methods
Fundamental and applied biological sciences. Psychology
Fundamental immunology
Immunity (Disease)
Molecular immunology
Molecular Structure
Pyrenes - chemistry
Rhodamines - chemistry
Techniques
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Description
Summary:A homogeneous noncompetitive immunoassay based on photoaffinity labeling techniques is described. Using this method, a fluorophore (reporter) can be specifically attached to an antibody in the vicinity of its antigen-combining sites. Upon antigen binding, changes in the fluorescence spectrum of the reporter molecule are often observed. Two fluorophores, pyrene and dansyl, were evaluated for this purpose. Also, this technology is ideal for fluorescence energy-transfer immunoassays that require labeling of the antibody with either a donor or acceptor fluorophore. In such cases, a fluorescent dye can be specifically attached near the antigen-combining site, where it can participate in high-efficiency energy transfer with its complementary fluorophore attached to the antigen.
ISSN:0003-2700
1520-6882
DOI:10.1021/ac00101a025