Fine specificities of murine anti-M-g monoclonal antibodies

The specificities of two murine anti-M-g monoclonal IgG1 antibodies, 3B10 a nd 2D5, were determined by pepscan analysis. The peptides which correspond to various fragments of amino-terminal portions of glycophorin A of group M (GPA-M), N (GPA-N) and Mg (GPA-M-g), and replacement analogues of some of these peptides, were synthesized on plastic pins and tested for binding of the antibodies. Both antibodies bound strongly to the N-terminal M-g octape ptide (1)LSTNEVAM(8), but they showed different subspecificities. The essen tial fragment of the epitope 2D5 are amino acid residues (STNEV6)-S-2. Repl acement of any of these amino acid residues by Ala, and replacement of Glu( 5) residue by Gly, abolished or strongly reduced the antibody binding, but replacement of Asn(4) by Thr gave only a moderate decrease of peptide activ ity. In contrast, the Leu(1) and Asn(4) residues were most essential compon ents of the epitope 3B10, while Ser(2), Thr(3) and Glu(5) seemed to be less important. Our present results and earlier ones on the specificity of huma n anti-M-g alloantibodies and monoclonal anti-M/M-g antibodies showed that antibodies reacting with M-g antigen recognize different fragments and/or d ifferent amino acid residues of the amino- terminal nonglycosylated domain of GPA-Mp. The knowledge of fie specificities of antibodies reacting with M -g antigen is interesting in view of the presence of anti-M-g alloantibodie s in 1-2% of human sera.