What Are the Basic Principles of the ELISA Assay Theory?


Quick Answer

ELISA, or enzyme-linked immunosorbent assay, measures the concentration of a factor called an analyte by attaching enzyme-linked antibodies to plastic plates via a substrate and observing their interaction with an antigen during incubation. The ELISA differs from other antibody assays in that by fixing the antibodies, it is possible to measure the analyte concentration. The analyte's presence and quantity indicate the presence or absence of the antigen.

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Full Answer

A single molecule attached to the antibody, also called antiglobulin, causes the substrate to change color upon detection of the antigen. The greater the color change, the higher the concentration of analyte. All ELISAs rely on the specific interaction between a small sequence of amino acids found on an antigen and a matching antibody binding site, requiring the use of both, although the order may differ. Either an antibody or an antigen are suitable for use in the first step of the ELISA, depending on the organism for which the laboratory is testing.

To perform a direct ELISA, the laboratory technician coats a plastic plate with the antibody or antigen. The technician then uses a protein-based solution to coat all unbound sites on the plate, adds the antigen or antibody to the plate, and then adds the substrate containing the linked enzyme. After the plate incubates, the technician then analyzes the enzyme-substrate reaction, which reveals whether the antigen is present, and if so, in what concentration.

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