An Efficient Method for Isolation of HL-A Antigens from Hematopoietic Cell Lines

Abstract

Abstract A simple isolation procedure has been established that gives a high yield of relatively pure HL-A antigens from cells of established hematopoietic cell lines. Particulate HL-A antigens were obtained from cells disrupted by shaking in buffer. These particulates were digested with papain in the presence of cysteine, and the soluble papain fragments that carried antigenic activity were purified by hypotonic dialysis, followed by ion exchange chromatography on DEAE-Sephadex, ultrafiltration, gel filtration and column electrophoresis. Several preparations of soluble HL-A antigens, each carrying one of the representative antigenic activities, HL-A1, HL-A2 or HL-A7, were obtained from three different cell lines. These preparations had a high degree of homogeneity in molecular size (48,000 Daltons) and a narrow range of electrophoretic mobility in the α2 globulin region. In these preparations, the overall recovery of HL-A antigenic activity was 30% to 37% for each isolated specificity, the recovery of OD280 units was 0.08% to 0.14%, the OD280/OD260 ratio was approximately 1.7 and the increase in specific antigenic activity was approximately 300 times that of papain digests.