ELUCIDATION OF MONOCLONAL-ANTIBODY POLYSPECIFICITY USING A SYNTHETIC COMBINATORIAL LIBRARY

The potential polyspecificity of an antipeptide monoclonal antibody wa s systematically examined using a soluble synthetic combinatorial libr ary (SCL). This SCL was composed of 400 different hexapeptide mixtures , each of which consisted of more than 130000 peptides totalling 50 mi llion individual sequences in approximately equimolar concentration. T he 400 peptide mixtures making lip this SCL were screened by competiti ve enzyme-linked immunosorbent as say (ELISA) for their ability to inh ibit monoclonal antibody binding to the original immunizing peptide. I ndividual peptides were derived front three different peptide mixtures of the peptide library through an iterative screening and selection p rocess; In addition to rile identification of the six-residue antigeni c determinant recognized by this monoclonal antibody two other hexapep tides were found to have binding affinities 5- to 10-fold higher than rite original antigenic determinant sequence. These peptide sequences represent analogs in which a polar amino acid (threonine) ill the orig inal antigenic determinant was substituted with a large, aromatic amin o acid (either tryptophan or tyrosine). Peptide analogs of the antigen ic determinant, ranging from single to multiple substitutions, as well as peptide sequences completely unrelated to the immunizing peptide, were also identified having binding affinities comparable to the origi nal immunogen. The present study illustrates the power of SCLs for the determination of alternative binding motifs Sor antigen antibody inte ractions. The use of SCLs in this manner inn? help to elucidate the ex tent of cross-reactivity poly-specificity and molecular mimicry found in antigen-antibody interactions.