Production of antibodies against murine class II antigens using unconjugated synthetic peptides of the β chain region 63-78

Synthetic peptides corresponding to residues 63-78 of the first domain of the β chain of murine I-A/I-E class II antigens were used in the unconjugated rather than the traditional protein-conjugated form to immunize (129J × B6)F1 mice. The sequences made represented the four haplotypes; A_β^k, A_β^d, A_β^bm-12 and E_β^d. These sequences were selected on the basis of computer algorithms used to predict surface accessibility and main-chain flexibility profiles, and by reported hypervriability and site-directed mutagenesis experiments of these regions. Factors such as the use of complete Freund's adjuvant, a continuous immunization regime, and the sex of the mice used were found to influence the amount of anti-peptide antibody produced when unconjugated peptide was used as the immunogen. Antibodies produced were shown by FACS analysis to react with I-A/I-E class II antigens expressed on the lymphocyte surface. These findings indicate that the use of unconjugated synthetic peptides representing sequences of proteins which are in close proximity to disulfide bonds may be an advantage over conventional methods of peptide conjugation to proteins.