Potency estimates of human gonadotrophins by a bioassay and three immuno-assay methods.

Abstract

ABSTRACT Human chorionic gonadotrophin (HCG), human menopausal gonadotrophin (HMG) and human hypophysial gonadotrophin (HHG) preparations were assayed by a bioassay and three immunoassay methods. The bioassay method was based on the increase in weight of the accessory reproductive organs of immature male rats. The immunoassays included haemagglutination inhibition, a radioimmunoassay using the double antibody technique and a solid phase radioimmunoassay using coated tubes. The same anti-HCG serum was employed in all immunoassays. When relatively crude HCG preparations (< 6000 IU/mg) were assayed, there was a highly significant (P < 0.001) correlation between the estimates obtained by all methods. However, when highly purified HCG preparations (> 6000 IU/mg) were assayed, the bioassay gave significantly higher potency estimates than the immunoassays. Furthermore, the estimates obtained by radioimmunoassays were significantly higher than those obtained by haemagglutination inhibition. However, the results obtained by the two radioimmunoassays showed a highly significant (P < 0.001) correlation. The regression coefficient of the line relating the logit transform of the response to loge, of the dose was significantly higher (b =−1.700±0.7322)) in assays employing the double antibody technique (a non-equilibrium system) than in solid phase (b =−0.945 ± 0.116) immunoassays (an equilibrium system). When highly purified HCG preparations were assayed by the double antibody technique, the regression coefficients were significantly higher than those obtained with the standard preparations. In relation to their biological activity, HMG preparations contained much less immunological activity, than HHG preparations. In addition, the regression lines obtained with HCG, HMG and HHG preparations showed significant deviations from parallelism when estimated by radioimmunoassays. The precision and sensitivity of the four assay methods increased in the following order: bioassay, haemagglutination inhibition, solid phase radioimmunoassay and radioimmunoassay involving the use of the double antibody technique. It is concluded that in relation to their biological activity, marked differences exist in the immunological activities of highly purified and relatively crude HCG preparations. In addition, the immunological activities of HCG, HMG and HHG preparations are dissimilar, when immunoassayed by the use of an anti-HCG serum.