Hypersecretion of the pituitary glycoprotein hormone alpha-subunit has been reported in pituitary adenomas, particularly in clinically nonfunctioning tumors and somatotroph adenomas. However, the prevalence of such hypersecretion has not been precisely defined. Using both a new highly sensitive and specific monoclonal antibody assay and a polyclonal antibody assay, serum levels of free alpha-subunit were compared in 63 unselected patients with these tumors, 19 patients with acromegaly, and 95 normal controls. In all patients the monoclonal assay detected a significantly greater number of subjects with elevated alpha-subunit levels than did the polyclonal assay (21 vs. 14; P less than 0.01). Fourteen of the 63 patients with clinically nonfunctioning tumors (22%) had elevated serum alpha-subunit levels in the monoclonal assay vs. 11 (17%) in the polyclonal assay. Among the 19 patients with acromegaly, the prevalence was 7 (37%) and 3 (16%) using the monoclonal and polyclonal assays, respectively. Twenty-eight (44%) of the patients with clinically nonfunctioning pituitary adenomas were female. Eleven (39%) of the women were under 45 yr old, as were 10 (29%) of the men. We conclude that the prevalence of free alpha-subunit hypersecretion in patients with clinically nonfunctioning and somatotroph adenomas may be higher than previously recognized, and that a sensitive and specific monoclonal antibody free alpha-subunit assay may provide a useful tumor marker in these patients. The prevalence of clinically nonfunctioning pituitary tumors among younger men and women may also have been previously under-estimated.