Production and characterization of a peptide specific, anti-major histocompatibility complex class II, monoclonal antibody

Monoclonal antibodies to major histocompatibility complex Class II proteins have been useful probes in understanding both the biochemistry and biology of these proteins. Almost all of the monoclonal antibodies previously described have been produced by immunization of mice with living cells. These antibodies react with native Class II proteins, but not usually with denatured material. It has been difficult to obtain specific anti-Class II antibodies which react with denatured proteins. Antibodies reactive with denatured proteins and with well-defined specificities would be useful in studies of Class II assembly and trafficking during the process of antigen presentation. In order to produce such an antibody we have immunized hamsters with a synthetic peptide corresponding to residues 146-177 (β 1 domain) of the mouse A^b_β protein. An antibody has been produced which reacts with the mouse Class II A_β chain from H-2^b, H-2^d, H-2^p, and H-2^q mice in immunoblotting assays, but not with the β chain from H-2^f, H-2^j, H-2^k or H-2^s mice. Comparison of the amino acid sequences of these proteins along with the reactivity patterns of the antibody on synthetic peptides corresponding to homologous regions from A^b_β, A^k_β, A^b_α and D^p suggest that the region of 153 to 155 is critical for the reactivity of this antibody. This antibody does not react with native Class II protein found on the surface of living mouse cells.