Electroimmunoassay of sex hormone binding globulin. Enhanced sensitivity by autoradiography using A-ring125I-17-β-oestradiol

Four different 125I-iodinated steroids were tested for their binding to human sex hormone binding globulin (SHBG) using an electrophoretic technique. 17-beta-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 electroimmunoassay of SHBG by adding the steroid to the samples before electroimmunoassay. 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 electroimmunoassay 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.