Endocrine: Measuring hormone levels

Most hormones are present in the circulating blood in extremely low concentrations, some as low as one millionth of a microgram (one picogram) per mL. This made measuring hormone concentrations difficult in early endocrinology. In the mid-20th century, however, a revolutionary technique called radioimmunoassay was developed, making it possible to measure hormones and their end products.

Radioimmunoassay occurs in several steps. First, an antibody specific for a hormone is mixed with a sample of purified hormone of known concentration that has been labeled with a radioactive isotope. Second, a sample of fluid from the organism to be tested is also added to this mixture. It is important to note that the amount of the antibody used has to be less than the amount needed to bind to the combined quantity of hormone in the fluid. This antibody deficiency creates competition among antibody binding sites (Step 3), and is the basis upon which the relative concentration of each sample can be determined.

In the final step, unbound antigens are removed from the mixture, and radioactive bound antigens are measured. Results of these experiments are compared to a “standard curve,” indicating the relationship between radioactive concentrations and organism hormone concentrations.

The method is summarized in Figure 1.

Figure 1 Radioimmunoassay; image adapted from Howell-Moroney, Madeleine, "Radioimmunoassay - Image". Embryo Project Encyclopedia ( 2022-01-24 ). ISSN: 1940-5030 https://hdl.handle.net/10776/13324

When compared with test assays, “standard curves” of hormone concentrations can elucidate the unknown concentrations within an error of 10-15 percent, allowing the assay of billionths or even trillionths of a gram of hormone.