A QUANTITATIVE ASSAY TO MEASURE THE INTERACTION BETWEEN IMMUNOGENIC PEPTIDES AND PURIFIED CLASS-I MAJOR HISTOCOMPATIBILITY COMPLEX-MOLECULES

A direct and sensitive biochemical assay to measure the interaction in solution between peptides and affinity-purified major histocompatibil ity complex (MHC) class I molecules has been generated. Specific bindi ng reflecting the known class I restriction of cytotoxic T cell respon ses was obtained. Adding an excess of beta(2)-microglobulin (beta(2)m) significantly increased the rate of peptide association, but it did n ot affect the rate of dissociation. Binding was complicated by a rapid and apparently irreversible loss of functional MHC class I at 37 degr ees C which might limit the life span of empty MHC class I thereby pre venting the inadvertent exchange of peptides at the target cell surfac e. All class I molecules tested bound peptides of the canonical octa- to nona-meric length. However, one class I molecule, K-k, also bound p eptides, which were much longer suggesting that the preference of clas s I molecules for short epitopes is not absolute and may be caused by factors other than the peptide-MHC class I binding event itself.