Human gamma globulin fractionation on anion exchange cellulose columns.

The serum proteins designated as y-globulins include all proteins of the slowest moving of the several major protein groups seen on serum electrophoresis conducted at alkaline pH (2). Among the y-globulins of normal serum are proteins of differing electrophoretic mobility (3,4), proteins with sedimentation coefficients of 6.6 S and 18 S in the ultracentrifuge (2, 3, 5, 6)) many individual antibodies (5, 7)) and the isohemagglutinins (8). In disease states, within the -y-globulins are also found the relatively specific serum properties identified in rheumatoid arthritis (9), Hashimoto’s thyroiditis (10, II), lupus erythematosus (12)) cold hemagglutination syndrome (13, 14)) multiple myeloma (15, 16)) macroglobulinemia (17) and cryoglobulinemia (1%. Study of the -y-globulins has been handicapped by the poor resolution of fractionation procedures and the amount of material or effort required to carry out each separation. The introduction by Peterson and Sober (19) of substituted cellulose ion exchangers with a high protein-binding capacity offered an opportunity to fractionate and characterize the y-globulins. Previous studies utilizing whole serum have demonstrated the capacity of anion exchange cellulose chromatography to separate the y-globulins into a number of regions of differing electrophoretic mobility (20, 21). However, with whole serum as a starting material, many of the chromatogram fractions containing y-globulins also included large amounts of other serum proteins. In the present work the y-globulins were first separated from whole serum by electrophoretic techniques and subsequently subfractionated by anion exchange cellulose chromatography. The y-globulin subfractions thus obtained have been characterized electrophoretically, ultracentrifugally, immunochemically, and by measurement of the hexose content. The findings demonstrate that the y-globulins are a heterogeneous group of proteins with differing physicochemical and immunological properties, and that anion exchange cellulose chromatography is a useful means of subdividing the y-globulins.