radioimmunoassay

A general method employing the reaction of antigen with specific antibody, permitting measurement of the concentration of virtually any substance of biologic interest, often with unparalleled sensitivity. The basis of the method is summarized in the competing reactions shown in the illustration. The unknown concentration of the antigenic substance in a sample is obtained by comparing its inhibitory effect on the binding of radioactively labeled antigen to a limited amount of specific antibody with the inhibitory effect of known standards.

Competing reactions that form basis of radioimmunoassay; * indicates the labeled antigen, and † “in known standard solutions or unknown samples.”

A typical radioimmunoassay is performed by the simultaneous preparation of a series of standard and unknown mixtures in test tubes, each containing identical concentrations of labeled antigen and specific antibody. After an appropriate reaction time the antibody-bound (B) and free (F) fractions of the labeled antigen are separated by one of a variety of techniques. The B/F ratios in the standards are plotted as a function of the concentration of unlabeled antigen (standard curve), and the unknown concentration of antigen is determined by comparing the observed B/F ratio with the standard curve.

The radioimmunoassay principle has found wide application in the measurement of a large and diverse group of substances in a variety of problems of clinical and biological interest. It is therefore not unexpected that there are differences in the specific methods employed for the assay of a particular substance. The full potential of the method has yet to be exploited. It seems that virtually any substance of biologic interest can be measured, the method being modified according to the characteristics of the particular substance. See also Antibody; Antigen; Immunology.

RIA (Royal Irish Academy), a learned body dedicated by its charter to ‘the cultivation of Science, Polite Literature, and Antiquities’; it came into being in 1785 with James Caulfield, Lord Charlemont, as its first President. The RIA was the immediate successor to the Hibernian Antiquarian Society, 1779-83, itself arising from the work of a Select Committee of the RDS, set up in 1772. In the 19th cent. it became the focus for philological, archaeological, and architectural studies. The RIA library became a major centre for the preservation of the literary and historical remains of Gaelic society. It now contains the Book of the Dun Cow, Leabhar Breac, the Book of Lecan, and an original autograph copy of part of the Annals of the Four Masters, along with 1, 400 manuscripts of various kinds.

A sensitive technique widely used to measure a range of biological substances, and to detect drugs such as peptide hormones and anabolic steroids during dope testing. The technique is based on the ability of an unlabelled form of the substance to inhibit competitively the binding of a radioactively labelled substance by specific antibodies. The concentration of the unknown sample is determined by comparing the degree of inhibition with that produced by a series of standards containing known amounts of the substance. The reliability of the test depends on the precise specification of the conditions under which the tests are made.

A sensitive assay method used for the measurement of minute quantities of specific antibodies or any antigen, such as hormones or drugs, against which specific antibodies can be raised; abbreviated RIA. An assay for a specific hormone uses antihormone antibody produced by injecting the human hormone into an animal, such as a rabbit. In the assay either the antigen or antibody is labeled with radioisotope.

Radioimmunoassay (RIA) is a very sensitive technique used to measure concentrations of antigens (for example, hormone levels in the blood) without the need to use a bioassay. It was developed by Rosalyn Yalow and Solomon Aaron Berson in the 1950s.^[1] In 1977, Rosalyn Sussman Yalow received the Nobel Prize in Medicine for the development of the RIA for insulin: the precise measurement of minute amounts of such a hormone was considered a breakthrough in endocrinology.

Although the RIA technique is extremely sensitive and extremely specific, it requires specialized equipment and is costly. It also requires special precautions, since radioactive substances are used. Therefore, today it has been largely supplanted by the ELISA method, where the antigen-antibody reaction is measured using colorimetric signals instead of a radioactive signal.

To perform a radioimmunoassay, a known quantity of an antigen is made radioactive, frequently by labeling it with gamma-radioactive isotopes of iodine attached to tyrosine. This radiolabeled antigen is then mixed with a known amount of antibody for that antigen, and as a result, the two chemically bind to one another. Then, a sample of serum from a patient containing an unknown quantity of that same antigen is added. This causes the unlabeled (or "cold") antigen from the serum to compete with the radiolabeled antigen ("hot") for antibody binding sites.

As the concentration of "cold" antigen is increased, more of it binds to the antibody, displacing the radiolabeled variant, and reducing the ratio of antibody-bound radiolabeled antigen to free radiolabeled antigen. The bound antigens are then separated from the unbound ones, and the radioactivity of the free antigen remaining in the supernatant is measured. Using known standards, a binding curve can then be generated which allows the amount of antigen in the patient's serum to be derived.

With this technique, separating bound from unbound antigen is crucial. Initially, the method of separation employed was the use of a second "anti-antibody" directed against the first for precipitation and centrifugation. The use of charcoal suspension for precipitation was extended but replaced later by Drs. Werner and Acebedo at Columbia University for RIA of T3 and T4.^[2] An ultramicro RIA for human TSH was published in BBRC (1975) by Drs. Acebedo, Hayek et al.^[3]

References

1. ^ Yalow RS, Berson SA. Immunoassay of endogenous plasma insulin in man. J Clin Invest 1960;39:1157-75. PMID 13846364.
2. ^ Werner SC, Acebedo G, Radichevich I (1974). "Rapid radioimmunoassay for both T4 and T3 in the same sample of human serum". J. Clin. Endocrinol. Metab. 38 (3): 493–5. PMID 4815178. 
3. ^ Acebedo G, Hayek A, Klegerman M, Crolla L, Bermes E, Brooks M (1975). "A rapid ultramicro radioimmunoassay for human thyrotropin". Biochem. Biophys. Res. Commun. 65 (2): 449–56. doi:10.1016/S0006-291X(75)80168-9. PMID 1148002. 

External links

• MeSH Radioimmunoassay
• Radioimmunoassay- procedure
• http://www.lib.mcg.edu/edu/esimmuno/ch4/radassay.htm

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