In vivo and in vitro androgen binding was studied in kidneys from male, female, and androgen-intensive (tfm/y) mice using sucrose gradient and dextran-coated charcoal assays. The mouse kidney differs from prostate in that it has very low 5a-reductase activity and, as a consequence, very little testosterone is converted to dihydrotestosterone. Following iv administration of 3H-testosterone, the unmetabolized steriod was bound by a 7.9S cytoplasmic and a 3.6S nuclear macromolecule. In vitrostudies indicated that this testosterone “receptor” complex in cytosol from castrated males hada Kd of 1.7 × 1O-9M binding 5.6 × 10-14 moles/mg cytosol protein. Similar values were obtained using cytosol from intact males and females. The protein nature and heat lability of the receptor and the need for cysteine groups for binding activity was demonstrated. The cytosol receptoralso bound 3H-dihydrotestosterone but with an apparent lesser affinity than for testosterone. Testosterone, dihydrotestosterone, androstenedione and progesterone inhibited 3H-testosterone binding with decreasing effectiveness. Metabolism studies at 1 C indicated that these steriods with the exception of testosterone, were rapidly metabolized by kidney cytosol preventing the accurate measurement of binding affinities for steroids other than testosterone. Estradiol was also found to bind to a 7.9S protein and could inhibit 3-H-testosterone binding. It is concluded that kidneys from male and female mice contain receptors with high affinity for testosterone per se. By contrast, there was no evidence of high affinity androgen binding by kidney cytosol from androgen-insensitive tfm/y animals (Endocrinology94: 746, 1974)