Proteins pp 349-355 | Cite as

Enzyme-Linked Immunosorbant Assay (ELISA)

  • Wim Gaastra
Part of the Methods in Molecular Biology™ book series (MIMB, volume 1)


In general, immunological methods are not very well suited for a quantitative determination of the antigen to be studied. The ELISA technique, however, can be used for a quantitative or at least semiquantitative determination of the concentration of a certain antigen. The method was first introduced by Engvall and Perlmann (1). The principle of ELISA (see Fig. 1), also called the double antibody sandwich technique, is the following: Antibodies against the antigen to be measured are adsorbed to a solid support, in most cases a polystyrene microtiter plate. After coating the support with antibody and washing, the antigen is added and will bind to the adsorbed antibodies. Next, a conjugate that will also bind to the antigen is added. Conjugates are antibody molecules to which an enzyme is covalently bound.
Fig. 1.

The main steps in a (noncompetitive) ELISA test. (1) The antibody to the antigen being quantitated is adsorbed onto a solid phase, usually polystyrene. (2) The sample containing the antigen being measured is then added. (3) Following the incubation and washing steps, a second enzyme-labeled antibody is then added. After further incubation and washing steps, enzyme substrate is added. (A substrate is chosen that will give a colored product). The amount of color produced is therefore proportional to the amount of antigen bound to the original antibody.


Substrate Solution Caprylic Acid Conjugate Solution Sodium Carbonate Buffer Polystyrene Microtiter Plate 
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  1. 1.
    Engvall, E., and Perlmann, P. (1971) Immunochemistry 8, 871–874.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press 1984

Authors and Affiliations

  • Wim Gaastra
    • 1
  1. 1.Department of MicrobiologyThe Technical University of DenmarkLyngbyDenmark

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