Progestins assay

Assay Methods for Evaluating Progestin Agonists and Antagonists. 

Steroid hormones achieve their effects on target tissues through intracellular receptor proteins. According to recent views, oestrogen and progestin receptors are localized in the nuclear compartment of the cells, whereas glucocorticoid receptors may reside in both the cytoplasm and the nucleus. Determination of the intracellular localization of androgen receptors awaits the development of (monoclonal) antibodies which will enable immunohistochemical studies. The molecular aspects of the mechanism of action of steroid hormones will be covered in other chapters [1-3] in this volume. The present chapter deals with the characterization, assay and purification of steroid receptors. 

Fig. 1. Assay for progestin receptors in human breast cancer cytosol. A. Saturation analysis. Total ( — ) and aspecific (o—o) binding are measured Specific binding (solid line) is calculated by subtraction. B. Scatchard plot derived from the binding data by the method of Chamness and McGuire [8]. Numerical data protein concentration of cytosol. 1.8 mg/ml assay volume, 100 /d cytosol volume, 50 /ul. Calculated values Kd = 0.76 nmol/l progestin receptor content, 510 fmol/mg protein.

Progesterone induces characteristic morphological changes of the endometrium after estrogen-induced proliferation of this tissue. This effect is the basis of the endometrium transformation assay, in which immature rabbits are primed with estradiol for several days and are subsequently treated with a progestin, either by oral application or by subcutaneous injection. The proliferating response of the uterus is examined by histological analysis and quantitated with the McPhail index on a scale from 1 to 4, with 4 as maximal effect. The assay is very sensitive and specific, even if the rabbit expresses only the B isoform of PR. 

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