Electroimmunoassay of sex hormone binding globulin. Enhanced sensitivity by autoradiography using A-ring ¹²⁵I-17-β-oestradiol

Abstract

Four different ¹²⁵I-iodinated steroids were tested for their binding to human sex hormone binding globulin (SHBG) using an electrophoretic technique. 17-β-oestradiol iodinated in its A-ring was found to bind with high affinity to SHBG. This radioactive steroid was used to increase the sensitivity of the electro-immunoassay of SHBG by adding the steroid to the samples before electro-immunoassay. The radioactive steroid incorporated into the immunoprecipitates could be observed by autoradiography.

The sensitivity of the assay, which employed a rabbit antiserum against purified human SHBG and was standardized with pure SHBG, was about 0.2 mg/l. The precision of the method, calculated as the coefficient of variation within and between assays, was 2.4% and 2.6%, respectively, for values within the normal range.

The mean SHBG concentration in healthy regularly menstruating women (n = 50) was 3.50±0.74 (SD) mg/l when measured in plasma, and 3.78±0.80 mg/l when measured in serum. The corresponding mean concentrations in healthy men (n = 28) were 2.26±0.45 mg/l in plasma and 2.44±0.49 mg/l in serum.

The modified eiectroimmunoassay described in this paper is a simple modification, which increases the sensitivity sufficiently to permit reliable quantification of SHBG over the entire range of concentration which could be relevant in clinical practice.

Key Words:

• autoradiography
• electrophoresis
• ¹²⁵I-iodinated steroids
• plasma protein
• reference values