The uptake of [1,2,4,5,6,7-3H]dihydrotestosterone into whole cells and nuclei has been assessed in fibroblasts grown from genital skin of 10 controls and 9 subjects with hereditary male pseudohermaphroditism due to androgen resistance. The cells were exposed to hypotonic buffer and ruptured by passage through a 25-gauge needle, and the nuclei were purified by sedimentation through 2.1 M sucrose. Uptake of the hormone into nuclei reached an apparent plateau in 45 min, was saturable at 1 nM dihydrotestosterone, and was not detectable in the presence of excess nonradioactive hormones. Over a wide range of uptake by intact cells from control subjects and from subjects with androgen resistance due to 5alpha-reductase deficiency or receptor deficiency, nuclear uptake averaged about half of the total cell uptake. Furthermore, in cells from two unrelated 46,XY phenotypic females with androgen resistance but normal 5alpha-reductase activity and normal whole cell dihydrotestosterone binding, uptake into the nucleus was also normal. In these subjects, the defect in androgen action must be at some terminal phase of androgen action.