Androgen- and estrogen-binding macromolecules from the hypothalamus plus preoptic area of 3- to 4-week-old mice have been detected and partially characterized. These components bind the respective hormones with high affinity (saturating at 4-8 nM) and sediment with rates typical of presumed steroid receptors (4.0-4.5 S in 0.15 M NaCl, 5.0-7.5 S without salt). A 90-95% reduction in androgen binding found in the androgen-insensitivity mutant mouse, testicular feminization (Tfm), provides a genetic control for the specificity of binding. This reduced androgen binding with Tfm/Y mutants and blocking experiments with non-radioactive estradiol [estra-1,3,5(10)-triene-3,17beta-diol] and testosterone (17beta-hydroxy-4-androsten-3-one) indicate the existence of at least two binding components: one with high affinity only for estradiol, the other with affinity for both androgens and estrogen. Based on these properties, a receptor mechanism that detects relative concentrations of androgens and estrogens is proposed.