The mixing of two histoincompatible human lymphocyte cell lines generated the release of a soluble factor which was capable of nonspecifically enhancing the in vitro immune response of normal mouse spleen cells against sheep erythrocytes. When active supernates were subjected to exclusion chromatography on Sephadex G-150 and BioGel P-150, the active principle eluted with molecules of approximately 35,000 mol wt. Column aliquots from similarly treated supernates from independent cultures of each lymphoid cell line were inactive. The human enhancing factor was concentrated and purified by ammonium sulfate fractionation, followed by Sephadex gel filtration and polyacrylamide gel electrophoresis.

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