A study of HLA (Bg) on red cells and platelets by immunoblotting with monoclonal antibodies

Abstract

HLA class I antigens (Bg) on red cells (RBCs) are expressed by some normal donors and by many patients with systemic lupus erythematosus (SLE). To identify the membrane components previously detected by hemagglutination with HLA class I-specific monoclonal antibodies (MoAbs), RBC membrane preparations were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotted with the HLA class I MoAbs. Two components were obtained that reacted with the MoAbs: a heavy chain of 45 kDa and a light chain termed beta2-microglobulin (beta2-M) of 11 kDa. The effect of chloroquine and acid elution in stripping HLA antigens is shown to be due to the removal of beta2-M, as only that component was detected in eluates from reactive RBCs. Neither antibody elution method affected the heavy chain expression assessed by immunoblotting. It is concluded that HLA class I antigens on RBCs are integral membrane components of the type normally found and wisely distributed on many nucleated cells. Platelets, which have stronger HLA class I antigen expression, were also studied, and their membrane preparations yielded heavy chain and beta2-M molecules; the effect of chloroquine treatment was harder to assess than that of acid elution, owing to the sensitivity with which both components are detected in immunoblotting. In eluates obtained from acid treatment only beta2-M is detected.