A rapid, sensitive and reliable assay for inhibin bioactivity.

Abstract

A rapid 2-day quantitative assay for inhibin bioactivity based on FSH secretion from pituitary cells of immature female rats is described. The bioassay exhibited steeper slopes, improved precision and greater (fourfold) sensitivity compared with a previously established pituitary FSH cell content assay. Whole pituitary glands were used for the preparation of pituitary cells and the method for cell dispersion required a single enzymatic treatment with trypsin. Cells (180,000 viable cells per well) were dispensed into culture media containing inhibin and incubated for 48 h. Media were removed and assayed for FSH by radioimmunoassay. Using a ram rete testis fluid preparation as standard the inhibin dose-response curves of 25 consecutive experiments showed indices of precision of -0.08(mean)[range -0.04 to -0.17] and Finney's G values of 0.017[0.003-0.06]. The mean ED40 was 0.17 units of inhibin activity per well with interassay variation of 16.2% at this point of the dose-response curve. The assay had a practical capacity of 400 wells, permitting the measurement of dose-response curves of at least 40 unknowns with three dose points and triplicate wells per dose. The assay is specific for inhibin-containing preparations from several animal species. Overall, the assay is simple, precise, and sensitive, indicative of its applicability to the measurement of inhibin samples with low inhibin bioactivity and to the screening of large numbers of fractions during inhibin purification.