In vivo activity of an H-2 alloantiserum purified by affinity chromatography on transplantation antigens

Transplantation antigens were isolated from murine tissue by solubilization with NP-40 and purification on a lentil lectin-Sepharose column. The glycoproteins eluted from the lectin column were coupled to CNBr-activated Sepharose to prepare a specific immunoadsorbent. Intact histocompatibility antigens were demonstrated on the carrier particles in a cytotoxic inhibition assay with specific antiserum and spleen lymphocytes. Adsorption and subsequent elution of alloantiserum directed against the coupled antigens yielded a 30-fold purified product. The presence of both active K and I region products on the column was demonstrated by the in vivo activities of the alloantibodies eluted from the coupled molecules. Both acute rejection, a function of H-2K antibodies, and passive enhancement, a function of Ia antibodies, could be induced by administration of the eluate to recipients of skin allografts. These in vivo results show that the adsorbent can be used for preparative purposes.