Evaluation of a radioimmunoassay for testosterone estimation

A radioimmunoassay technique, which is essentially a modification of the method described by Furuyama et al.[1], has been evaluated for the determination of testosterone in human peripheral plasma and rat testis tissue.The antiserum used was raised against testosterone-3-(0-carboxymethyl)-oxime-bovine serum albumin in female rabbits. It had an association constant of 5.5 × 10⁹ 1/mol, 4°C, at a dilution of 1 in 20,000. The procedure involved addition of [³H]-testosterone internal standard, extraction and chromatography of the plasma extracts on alumina micro-columns prior to assay. Testis tissue extracts were not chromatographed. Known amounts of standard testosterone were subjected to the same procedures.After incubation with antiserum for 16 h at 4°C total recovery from the extraction, chromatography (when used) and incubation procedures were measured in order to correct for losses. Either toluene scintillation fluid, dextran-coated charcoal or polyethylene glycol were used to separate free and bound testosterone. For human plasma as well as for testis tissue a good correlation was observed between results obtained with radioimmunoassay and a gas chromatographic method using electron capture detection of testosterone chloroacetate[12].