Assay of human endometrial progesterone receptors using the natural hormone and a polyethylene glycol precipitation technique

Abstract

An accurate, reliable assay for progesterone receptors of human endometrium has been developed. After equilibration of tissue extracts with tritium-labelled progesterone, bound steroid was precipitated using polyethylene glycol (PEG) in the presence of γ-globulin. The receptor-hormone complexes were not dissociated during the procedure and interference by nonspecific binding components, including corticosteroid binding globulin, was corrected for by equilibration after mild heat treatment. The presence of a 50-fold excess of cortisol during equilibration had little effect on binding site concentrations but gave rise to a decrease in the dissociation constants ( K D ); a 500-fold excess reduced both the receptor site concentrations and K D values. Glycerol (10%), present during equilibration only, had little effect on either the number of receptor binding sites or affinity. Comparison of the PEG method with a published method using dextran-coated charcoal and the natural hormone showed differences that may perhaps be explained by variable dissociation of progesterone from nonspecific and possibly specific binding components in the presence of charcoal. Binding site concentrations and K D values were of a similar order when using either progesterone or the synthetic analogue, R5020. R5020, however, tended to give erratic results more often than progesterone.