ⓘ ELISA

                                     

ⓘ ELISA

The enzyme-linked immunosorbent assay is a commonly used analytical biochemistry assay, first described by Engvall and Perlmann in 1971. The assay uses a solid-phase enzyme immunoassay to detect the presence of a ligand in a liquid sample using antibodies directed against the protein to be measured. ELISA has been used as a diagnostic tool in medicine, plant pathology, and biotechnology, as well as a quality control check in various industries.

In the most simple form of ELISA, the antigen from the sample are attached to the surface. Then, the appropriate antibody, which is applied to the surface so it can bind to the antigen. This antibody is linked to an enzyme at the final step, a substance containing the enzymes substrate is added. The subsequent reaction gives a recognizable signal, most commonly a color change.

Performing ELISA involves at least one antibody with specific antigen. A sample with an unknown amount of antigen is immobilized on a solid support, usually polystyrene microtiterplate plate or not specifically through adsorption on the surface or specifically via capture by another antibody specific to the same antigen, in a "sandwich" ELISA. After the antigen is immobilized, the detection antibody is added, forming a complex with the antigen. The detection antibody can be covalently linked to an enzyme or can itself be detected by a secondary antibody that binds to an enzyme through conjugation. Between each step the plate is typically washed with a weak detergent solution to remove any proteins or antibodies that are not particularly related. After the final wash step the plate is developed by adding an enzymatic substrate to produce a visible signal, which indicates the quantity of antigen in the sample.

It should be noted that ELISA can perform other forms of ligand binding assays and not strictly "immuno" assays, though the name carried the original "immuno" because of the common use and history of development of this method. The technique essentially requires any ligating reagent that can be immobilized on the solid phase along with a detection reagent that will bind specifically and use an enzyme to generate a signal that can be correctly identified. Between the washes, only the ligand and its specific binding counterparts remain specifically bound or "immunosorbed" by the interaction of antigen-antibody on the solid phase, while the nonspecific or unbound components are washed away. Unlike other spectrophotometric lab assay formats where the same reaction well, for example, cuvettes can be used repeatedly after washing, the ELISA plates have the reaction products immunosorbed on the solid phase, which is the part of the plate, and so are not easily reusable.

                                     
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