A patient with hypopituitarism and an unusually large chromophobe adenoma which secreted an enormous amount of human PRL (hPRL) provided us with a unique opportunity to study the heterogeneity of hPRL. Serum samples obtained before and after surgery as well as pituitary extract were studied. Each sample contained three distinct hPRL peaks on gel filtration designated as big medium, and small. The hPRL moiety of each peak fraction was stable in size, immunoreactivity, and bioreactivity, indicating that the polymorphic hPRL components, once formed, were not interconvertible. As the relative proportion of medium and small hPRL components in all blood samples obtained before surgery and in the pituitary tumor extracts were similar, it seems that these two forms of hPRL originate from the tumor. However, there was a lack of correlation between big, medium, and small hPRL in samples obtained before and 1 yr after surgery. Also, increasing amounts of radioactive substance similar to increasing amounts of radioactive substance similar to big PRL were formed by exposure of 125I-labeled small PRL to progressively larger concentrations of serum. Thus, only small and possibly medium PRL are secretory products.