Beta-2-Microglobulin of Rat Major Histocompatibility Complex Class I Antigens

Abstract

Detection of the β₂-microglobulin (β₂m) component of the rat MHC class I antigens has been difficult. In the present report, we have addressed this issue by a systematic study of rat class I antigens from red blood cells or from lymphocytes that were freshly isolated or cultured in the presence of autologous or heterologous sera and surface-labeled with ¹²⁵I or intrinsically labeled with radioactive amino acids. First, specific radioiodination of rat β₂m in association with the antigen heavy chain on red blood cells or lymphocytes is minimal, resulting in its poor identification by SDS-PAGE. Second, labeling with radioactive methionine or lysine gives a more intense β₂m band with respect to the heavy chain than labeling with arginine or tyrosine. Third, the β₂m component shows a large increase in intensity compared to the heavy chain when the antigens are isolated from lymphocytes that are cultured in the presence of fetal bovine serum prior to ¹²⁵I-labeling. This increase is due to exchange of endogenous rat β₂m with bovine β₂m and to a much higher level of radioiodination of the latter. Fourth, rat red blood cells and lymphocytes contain free surface β₂m molecules in addition to those associated with the antigen heavy chains. The free molecules show a much higher level of radioiodination than those associated with the heavy chains, and there is little exchange between the antigen-bound and the free β₂m after radioiodination. Thus, intrinsic labeling with suitable radioactive amino acids rather than surface labeling with ¹²⁵I is the method of choice for unambiguous detection of the endogenous β₂m component of the rat MHC class I antigens.