Serum levels of dihydrotestosterone in male rhesus monkeys estimated by a non-chromatographic radioimmunoassay method

Abstract

Since the antiserum used to measure dihydrotestosterone (DHT) has considerable cross-reaction with testosterone (T), the chromatographie separation of DHT from T is a pre-requisite to get precise estimates of DHT levels by radioimmunoassay (RIA) in biological fluids. To simplify this procedure a non-chromatographic method for the estimation of DHT is reported. The treatment of T with 0.5% aqueous solution of potassium permanganate (KMnO₄) for 30 min, at room temperature, selectively destroys its cross-reactivity with antiserum raised against T-3(O-carboxy-methyl)-oxime-bovine serum albumin. Whereas, the thin layer Chromatographie mobility of DHT in benzene-ethyl acetate solvent system (2:1, v/v) and its cross-reaction with T-antiserum remain unaffected (70%) following treatment with KMnO₄. Thus by treating the ether extracts of biological samples with KMnO₄ the endogenous T is inactivated and a simple non-chromatographic method is validated for quantitating DHT by RIA. In this assay procedure the serial dilutions of pools of serum samples from adult rhesus monkeys show dose-response curves that are parallel to the DHT standard curve. The correlation coefficient of DHT estimates obtained after Chromatographie separation of DHT on sephadex LH-20 column, using cyclohexane-benzene-methanol (80:15:15, by vol.) solvent system, or following KMnO₄ treatment of ethereal extracts of serum samples is 0.99. Using this method, the geometric mean serum levels of DHT. measured in 20 adult male rhesus monkeys, are significantly (P < 0.05) higher at night (5.38 nmol/l) than during the day (3.22 nmol/l).