The effect of testosterone on regulation of epididymal protein synthesis has been investigated in castrated rhesus monkeys (Macaca mulatta). The proteins in the treated monkeys were characterized using polyacrylamide gel electrophoresis (under nondenaturing and denaturing conditions) and electrofocusing. At least four distinct proteins have been shown to be synthesized by the monkey epididymis under testosterone influence. Two of these proteins were detected following two days of testosterone treatment while the other two proteins were detected after a six-day treatment period. None of these proteins was detectable in monkeys treated with estradiol for six days. Electrofocusing of epididymal cytosol proteins from untreated and testosterone-treated and castrated monkeys also confirmed the presence of four androgen-dependent proteins in this species. The isoelectric points of these proteins were shown to range between 5.8 and 6.4. The molecular weights of these proteins were found to vary between 47,500 and 66,000. The in vitro incorporation of 3H-labeled amino acids was markedly greater in the androgen-primed epididymis as compared with the control tissue.