Inhibin is a glycoprotein hormone that is defined on the basis of inhibition of pituitary FSH production, However, previous data have not shown any correlation between RIA measurements of inhibin and FSH in men. New enzyme-linked immunosorbent assays, specific for inhibin A, inhibin B, and inhibin pro-alphaC-related immunoreactivity, were applied to the measurement of inhibin in 32 healthy men. Further measurements of inhibin B and pro-alphaC-RI were carried out on groups of men exhibiting a wide range of FSH concentrations, including semen donors, infertile men, and men with elevated FSH concentrations. Inhibin A was undetectable (<2 pg/mL) in all men studied. The healthy men studied all had measurable concentrations of inhibin B (135.6 pg/mL; confidence interval, 108.4-169.4) and pro-alphaC-RI (426.3 pg/mL; confidence interval, 378.4-480.2). A close negative correlation was found between the inhibin B and FSH concentrations in the semen donors (r = -0.69; P < 0.001), the infertile men (r = -0.81; P < 0.001), and the men with elevated FSH concentrations (r = -0.54; P < 0.01), but not in a group of healthy volunteers (r = -0.08; P = NS). No correlation was observed between concentrations of pro-alphaC-RI and FSH in any of the groups studied. These results strongly suggest that the physiologically important form of inhibin in men is inhibin B, which has a critical effect on FSH release. Inhibin B may offer a clinically useful serum marker of testicular function.