MEASUREMENT OF FREE AND OCCUPIED CYTOPLASMIC AND NUCLEAR ANDROGEN RECEPTOR SITES IN RAT VENTRAL PROSTATE GLAND

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Abstract
SUMMARY: A method has been developed which allows the estimation of occupied and unoccupied androgen receptor sites in both cytoplasmic and nuclear fractions of rat ventral prostate. The procedure involves precipitation of receptor proteins and incubation of precipitates with labelled 5α-dihydrotestosterone. Uptake of 3H-labelled steroid at 0–4 °C gives an indication of free receptor, whereas binding at a raised temperature (15 °C) allows estimation of occupied receptor. Non-specific binding was measured in the presence of a 100-fold excess of unlabelled 5α-dihydrotestosterone. The exchange method was specific for androgens, and specific binding was detected only in fractions of androgen-dependent tissues. The method can be applied to cytosol, whole nuclei, chromatin and salt-extractable and salt-resistant protein preparations from nuclear fractions, and gives a reliable estimate of total receptor sites when occupied as compared with control measurements of unoccupied sites.