ISOLATION OF HUMAN PITUITARY PROLACTIN

A process developed earlier for the extraction of human follitropin, lutropin, thyrotropin and growth hormone from homogenizeed frozen pituitaries provided a residue utilized for the isolation of prolactin. The isolation procedure involved extraction at pH 9.8, molecular sieve chromatography on Sepharose CL-6B, hydrophobic interaction chromatography on phenyl-Sepharose CL-4B, molecular sieve chromatography on Sephadex G-100 Superfine, and ion-exchange chromatography on DEAE-Sepharose CL-6B using a convex gradient. The progresive purification was guided by radioimunoassays. The final product was obtained in yields of 31 μg/gland, and was equipotent with a pituitary preparation (VLS-3) supplied by the National Pituitary Agency (NIH, Bethesda, U.S.A.). Contamination by growth hormone was low (less than 2%), and by other pituitary protein hormones negligible (less than 0.05%). No heterogeneity of the isolated prolactin was observed by sedimentation-equilibrium analysis in the ultracentrifuge, by SDS electrophoresis in polyacrylamide gel or by molecular sieve chromatography in 6 M guanidine hydrochloride. These different techniques gave values in the range of 21 000-23 000 for the molecular weight of prolactin. In free zone electrophoresis, and also in polyacrylamide gel electrophores is the prolactin preparation was, however, heterogenous and resolved at alkaline pH into three distinct components. The former technique permitted isolation and assay of the components, indicating that they were all fully active. © 1979.