Radioimmunological characterization of human thyrotropin and its subunits: applications for the measurement of human TSH

ABSTRACT. Radioimmunological studies were carried out with antisera to the alpha- and betasubunits of human thyrotropin (hTSH) and to intact hTSH. In the homologous hTSH-α assay (anti-hTSH-α and 125I hTSH-α), immunoreactivity of hCG-α was the same as that of hTSH-α. The dose-response curves of intact hTSH, hLH, hFSH, and hCG, though showing less inhibition than the alpha subunit, were parallel in this assay, thus reflecting structural similarity between the alpha subunit of hTSH and the other glycoprotein hormones. Bovine TSH-α was essentially unreactive in the homologous human alpha assay. In the homologous hTSH-β assay (anti-hTSH-β and 125I hTSH-β), the immunological specificity of the beta subunit was confirmed. Intact hCG and human menopausal gonadotropin did not crossreact; however, a low degree of cross reactivity of intact hFSH and intact hLH was observed which may be explained by contamination with small amounts of hTSH. There was a low but definite cross reactivity against bTSH-β in confirmation of studies by double diffusion. No common antigenic sites were detected in the two subunits against either anti-hTSH-α or anti-hTSH-β; the small percent cross reaction of hTSH-α in the beta-unit assay and of hTSH-β in the alpha-unit assay, we concluded was caused by contamination of the β-unit with α-subunit and vice versa. This conclusion is based upon the identity of the hCG-α and hTSH-α in the alpha assay, and their greatly different reactions in the beta assay. From these studies it appears that the conformations or, at least, the immunological determinants of the two subunits available for reaction with antibody are somewhat different than in the intact hormone. Thus, the affinity of intact hTSH and the slope of its reaction compared to hTSH-β were different, both in the homologous anti-hTSH system and the homologous anti-hTSH-β system. Nevertheless, the affinity of hTSH and hTSH-β for hTSH-β antiserum were quite similar if intact hTSH was used as the label. Under these conditions, a specific and sensitive hTSH assay could be developed with antisera to hTSH-β. However, its sensitivity and specificity were not greater than the best homologous hTSH assays reported.