What is a Serum ELISA?

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  • Written By: S. Gadd
  • Edited By: J.T. Gale
  • Last Modified Date: 10 November 2019
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The serum enzyme-linked immunosorbent assay (serum ELISA) is a method used to determine the amount of a biological substance in a serum sample by relying on the specific interaction between an antigen and an antibody. Examples of biological substances of interest include an antibody produced in response to a viral infection, such as an antibody against human immunodeficiency virus (HIV), or to a hormone that indicates pregnancy, such as human chorionic gonadotropin (hCG), or to an autoimmune antibody produced in rheumatoid arthritis, such as rheumatoid factor. In general, there is a direct assay, which uses a specific antibody to detect the presence of antigen in a serum sample, and an indirect assay, which uses an antigen to determine the presence of antibodies in a serum sample.


In the direct serum ELISA, a sample containing an unknown amount of antigen is attached to an immobile surface, such as a reaction tube or a microtiter plate, and is incubated with a specific antibody that is chemically linked to an enzyme. Conversely, in the indirect serum ELISA, a known antigen is attached to an immobile surface and then is incubated with the serum sample that contains an unknown amount of antibody. The antigen-specific antibody in the serum sample is expected to bind tightly to the immobilized antigen, while non-specific antibodies are removed in the following wash steps. Generally, the immobile surface is treated with a second antibody that recognizes the unvarying region of all antibodies, and which is chemically linked to an enzyme. In both the direct and indirect ELISA, the final step consists of addition of an enzyme-specific substrate, initiating a reaction that produces a measurable signal that is directly proportional to the amount of antigen or antibody present in the serum sample.

The serum ELISA is a widely used test for several reasons. Most importantly, it is considered a reliable clinical test due to the specificity of the antigen-antibody interaction, and the sensitivity of this assay allows detection of biological substances at extremely low concentrations in serum. It is designed for the assessment of a large number of samples at the same time, and so is often used in large-scale operations, such as the screening of blood donor samples for the presence of HIV antibodies. In addition, ELISA kits that measure commonly assayed antibodies and antigens are commercially available for use in clinical or research settings, and conveniently contain all necessary reagents for carrying out a full experiment.

A growing area of study involves the application of serum ELISA technology in the area of cytokine detection. Cytokines are soluble protein molecules secreted by the immune system that are often involved in inflammatory processes; cytokine levels are therefore informative regarding disease states associated with chronic inflammation, such as heart disease, autoimmune diseases, and digestive diseases. It is thought that assessment of cytokine levels may be able to distinguish between inflammatory bowel diseases, such as Crohn’s disease and ulcerative colitis, and may eventually be able to predict outcome in heart disease and rheumatoid arthritis.


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