Quantitation of antigens by enzyme immunoassays

A number of procedures can be used for the precise measurement, by enzyme immunoassay, of minute amounts of antigens. Enzyme immunoassays, or in their abbreviated form EIA, are divided into homogeneous and heterogeneous tests. In homogeneous assays there is no need to separate the antigen-antibody immune complexes from free antigen and/or free antibody; however, such a separation is required in heterogeneous assays. The heterogeneous enzyme immunoassays are described in the entry-entitled ELISA.

The homogeneous assays are based on antibody-mediated changes in enzyme activity. A specific antibody, by reacting with an antigen-enzyme conjugate, diminishes the capacity of the enzyme to interact with its substrate and, consequently, decreases the enzyme's activity; addition of free antigen proportionally restores the enzyme's activity. Satisfactory results are obtained in these assays only when the antigen is hapten and the enzyme label is cither glucose-6-phosphate dehydrogenase or malate dehydrogenase. To date, the most widely used homogeneous enzyme immunoassay, known commercially as EMIT (enzyme-multiplied immunotechnique), is employed, essentially to test drugs. It is rapid and simple. However, its sensitivity is limited and therefore it is applicable only when ligands are present in relatively high concentrations. Several variants of this basic homogeneous enzyme immunoassay employing enzyme inhibitors, prosthetic enzyme groups, enzyme substrates and two enzymes catalyzing consecutive reactions have been described. Although they are based on original and interesting principles, such procedures are only rarely employed, if at all.

See also: Electron microscopy, immunological applications; Enzyme-linked immunosorbent assay (ELISA); Immunoassays; Immunocytochemistry and enzyme markers; Western blotting.

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