To determine whether glycosylated human PRL (G-hPRL) is a circulating form of hPRL, serum samples obtained from normal men and women were studied. The PRL was immunoprecipitated from 100-.mu.L aliquots of serum, and the precipitates were subjected to gel electrophoresis in the presence of sodium dodecyl sulfate, electrotransferred to nitrocellulose paper, immunoblotted with anti-hPRL serum and [125I]protein-A, and autoradiographed. The autoradiographs showed an intensely stained G-hPRL band, with a mol wt of 25K, in almost all samples studied. The nonglycosylated PRL (hPRL) band, with a mol wt of 23K, was much more variable in staining intensity from individual to individual. Some samples also had a 27K glycosylated immunoreactive PRL band. Serum samples were also obtained every other day from two women throughout a 28-day period; again, each sample had an intensely stained G-hPRL band and a hPRL band that varied in staining intensity from day to day. In contrast, however, to the G-hPRL/hPRL pattern in serum samples obtained from normal women and men, samples from pregnant women at the onset of spontaneous labor showed opposite results, that is an intensely stained hPRL band but only a faintly stained G-hPRL band. These results indicated that G-hPRL is an important component of circulating PRL in normal men and women and that it may vary in amount in certain physiologic states.