Radioimmunoassay: Principle, Uses, Limitations

Radioimmunoassay (RIA) is very sensitive (can detect at a concentration of <0.01 μg/mL) and a specific technique that is used for the quantitative detection of antigens or haptens.

RIA is an extremely important tool in biomedical research and clinical practice. It is commonly used to assay microbial antigens, proteins, vitamins, hormones, or drugs in serum. A specialized RIA called radioallergosorbent test (RAST) is used to measure the amount of serum IgE antibody that reacts with a known allergen (antigen).

Because of the radio hazard associated, the use of RIA is reduced and has been largely replaced by ELISA testing.

Radioimmunoassay was developed by Berson and Yalow (1960), for which they were awarded Nobel prize in 1977.

The principle of RIA is similar to that of competitive ELISA for antigen detection, except that nucleotides (usually ¹²⁵I or ¹⁴C) are substituted for enzymes in RIA.

Principle of Radioimmunoassay (RIA)

Radioimmunoassay uses radioisotope-labeled purified known antigen which competes with unlabeled (unknown) antigen for binding sites on a known amount of antibody.

The Ag-Ab complexes that form between the antigen and antibody can then be precipitated using the second antibody and the amount of radioactivity of the bound complex is measured by means of radioisotope analyzers and autoradiography.

When the sample contains a high amount of antigen, much of the antigen-binding sites of the antibodies are occupied by unlabelled antigen. So the bound complex will show little radioactivity. The radioactivity generated, in fact, is inversely proportional to the amount of antigen present in the sample.

Then the concentration of the unknown (unlabeled) antigen or hapten present in a sample is determined by plotting the value of radioactivity generated in a standard chart generated using different concentrations of same antigens against the radioactivity of the bound complex.

Advantages

1. Highly sensitive so it can be used for the detection of small quantities of analyte or antigens.
2. quantification of antigens or analytes is possible with RIA.

Limitations

1. The handling of radioisotopes requires specific safety measures. Because of this limitation, RIA has been replaced by ELISA in clinical laboratories.

References and further readings

1. Madigan Michael T, Bender, Kelly S, Buckley, Daniel H, Sattley, W. Matthew, & Stahl, David A. (2018). Brock Biology of Microorganisms (15th Edition). Pearson.
2. Willey, Joanne M, Sherwood, Linda M, & Woolverton, Christopher J. (2016). Prescott’s Microbiology (10 edition). McGraw-Hill Education.
3. Tille, P. (2017). Bailey & Scott’s Diagnostic Microbiology (14 edition). Mosby.