Enzyme-Linked Immunosorbent Assay Principle And Elisa Test Procedures

Enzyme-linked immunosorbent assay (ELISA) is also known as an enzyme immunoassay (EIA). ELISA is defined as a biochemical technique used in many applications, including microbiology, blood screening, veterinary, and immunology, to detect antigens and antibodies present in a sample.

ELISA technology is considered an important diagnostic tool in plant pathology and medicine. It is also used for quality control in the food industry. To know more about ELISA, you can also visit www.bosterbio.com/human-granzyme-b-picokine-trade-elisa-kit-ek1114-boster.html.

ELISA relies on specific antibodies to fix the target antigen. The detection system is then used to detect the presence and amount of antigen-binding.

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Enzyme-Linked Immunosorbent Assay (ELISA) uses various antigen-antibody combinations, including always antibody or enzyme-labeled antigen, and enzyme activity is calculated colorimetrically.

Enzyme activity was measured using a substrate that changes color when modified by an enzyme. The light absorption of the product formed after the addition of the substrate is measured and converted into a numerical value. Depending on the antigen-antibody combination, different types of ELISA are referred to as direct ELISA, indirect ELISA, sandwich ELISA, competitive ELISA, and others.

The reagents in the ELISA assay are immobilized, which facilitates the execution of the procedure. The assay includes an antibody layer on the microtiter plate.

A popular antibody is IgG, which is purified and used in conjugates to avoid interference from other proteins when bound to enzymes. When a blood sample is added, the correct antibody (primary antibody) attaches to the protein of interest (eg a cytokine).

The secondary antibody binds to a unique epitope on the protein. The assay is labeled with biotin, which allows subsequent binding of proteins such as streptavidin-conjugated enzymes. The enzymes that are often used during this process are peroxidase (HRP) and alkaline phosphatase (AP).

All unbound reagents/serum components were removed by washing the plaque thoroughly. PBS-T (Phosphate Buffered Saline with Tween) is used as a diluent to remove unbound molecules.