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Related Experiment Videos

A sensitive micro-immunoassay using beta-galactosidase/anti-beta-galactosidase complexes.

H Durbin, W F Bodmer

    Journal of Immunological Methods
    |February 26, 1987
    PubMed
    Summary

    A novel microELISA assay utilizes enzyme/anti-enzyme complexes for highly sensitive antigen detection. This method is ideal for analyzing limited antigen amounts, such as histocompatibility antigens on lymphocytes.

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    Area of Science:

    • Immunology
    • Biochemistry
    • Analytical Chemistry

    Background:

    • MicroELISA assays are crucial for sensitive biological molecule detection.
    • Enzyme/anti-enzyme complexes offer enhanced signal amplification.
    • Unlabelled antibody bridging is a common immunoassay technique.

    Purpose of the Study:

    • To develop a highly sensitive microELISA assay.
    • To utilize enzyme/anti-enzyme complexes for improved detection limits.
    • To demonstrate the assay's utility in analyzing histocompatibility antigens.

    Main Methods:

    • A microELISA assay was developed using enzyme/anti-enzyme complexes.
    • Beta-galactosidase/anti-beta-galactosidase complexes were formed using monoclonal antibodies against bacterial beta-galactosidase.

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  • Enzyme activity was quantified using a fluorogenic substrate.
  • The assay was applied to analyze Class I and Class II histocompatibility antigens on peripheral blood lymphocytes.

    • Main Results:

    • The developed microELISA demonstrated high sensitivity.
    • The assay is suitable for detecting limited amounts of antigen.
    • Successful analysis of histocompatibility antigens on 5000 lymphocytes per well was achieved using Terasaki or microtitre plates.

    Conclusions:

    • The new microELISA assay provides a sensitive method for antigen detection.
    • This assay is particularly valuable for analyzing scarce antigens.
    • The method is applicable to the study of cell surface antigens like histocompatibility antigens.