Determination of Low Amounts of Specific Binding

1. An PHJoestradiol-exchange methodwas developed for the determination ofoestradiolreceptor complexes in the nuclear fraction of immatur rat testicular tissue. This method permits the determination of nuclear oestradiol-receptot sites in the presence of a relatively large amount of non-specific oestradiol binding present in testicular nuclei. After incubation of nuclei for 60min at 200C in the presence of EH]oestradiol with or without a 1000-fold excess of non-radioactive diethylstilboestrol, specific binding can be determined quantitatively in the KCI-extractable fraction, which contains 40% of the total receptor population. 2. The amount of receptor-bound steroid present in the 0.4M-KCI extract of testicular nuclei remained constant during incubation at 200C. For uterine nuclei incubated with r3Hloestradiol at 37°C a shift of specifically bound [3fHfoestradiol occurred from the KCI-soluble fraction to the KCI-isoluble fraction. 3. In intact rat testis, about 20% of the total receptor coneentration was present in its nuclear form. Hypophysectomy 5 days before measurement resulted in a twofold decrease in the amount of receptor, which was present mainly in the cytosol. After injection of choriogonadotropin to intact animals, the total receptor concentration increased threefold. 4. This nuclear exchange method might be useful for determination of occupied specific receptor sites in tissues with telatively low contents of specific receptors.