Isolation of a human B lymphocyte membrane protein with Ia-like properties

Abstract

A rapid method for isolation of a major surface membrane glycoprotein from whole, unfractionated cultured human B [bone marrow-derived] lymphoblasts is described. After detergent solubilization the method uses gel filtration followed by affinity chromatography on Sepharose Con [concanavalin] A and then alkaline acrylamide gel electrophoresis. Specific, high-titer, rabbit antisera to the isolated protein reacted with cultured and normal peripheral blood B lymphocytes and peritoneal macrophages from a renal dialysis patient. The antisera selectively inhibited the mixed lymphocyte reaction at high dilution. The protein reacted with a heterologous antiserum to HL-B antigens [Ag] and contained subunits of MW 33,000 and 27,000. Resolution of the subunits required a discontinuous SDS [sodium dodecyl sulfate] gel system. These properties indicate its similarity to murine Ia [immune response associated] Ag. The protein was not associated with .beta.2 microglobulin and showed no structural or antigenic similarity to the major erythrocyte glycoprotein, glycophorin. Antisera to the protein failed to precipitate surface-radioiodinated components from similarly treated extracts of cultured human T [thymus-derived] lymphoblasts. This method now makes available a reference membrane glycoprotein from a differentiated, nucleated human cell in sufficient purity and quantity for kinetic and biosynthetic studies.