COMPETITION INHIBITION OF CYTOTOXIC T-LYMPHOCYTE (CTL) LYSIS, A MORE SENSITIVE METHOD TO IDENTIFY CANDIDATE CTL EPITOPES THAN INDUCTION OF ANTIBODY-DETECTED MHC CLASS-I STABILIZATION

We compared the efficiency of two commonly used cellular major histoco mpatibility complex (MHC) class I peptide-binding assays to identify a cytotoxic T lymphocyte (CTL) epitope-containing peptide among length variants derived from the human papilloma virus type 16 (HPV 16) oncop rotein E7. Although both assays identified the same sequence (E7 49-57 ) as the most efficient D-b-binding peptide, the efficiency by which t hey did so differed markedly. In a peptide competition cytotoxicity (P CC) assay, based on inhibition of CTL lysis by competition for binding to MHC class-I molecules between a known CTL epitope-containing pepti de and peptide of interest, E7 49-57 bound 45-fold more efficiently to D-b than the second D-b-binding peptide in line. In the widely used R MA-S MHC class I peptide-binding assay, based on peptide-induced stabi lization of 'empty' MHC class-I molecules at the surface of antigen-pr ocessing defective RMA-S cells, this difference was only 3 fold. Simil ar differences were observed when other D-b-restricted CTL clones and CTL epitope-containing peptides were used in the PCC assay. The same p henomenon was observed when peptide binding affinities for H-2K(b) wer e analyzed in both assays. We conclude that the PCC assay discriminate s more efficiently between high- and low-affinity MHC class I binding peptides than the RMA-S assay. This observation is ascribed to the fac t that peptide-MHC class I dissociation is an important parameter in t he PCC but not the RMA-S assay.