CLASS-I MAJOR HISTOCOMPATIBILITY COMPLEX-RESTRICTED CYTOTOXIC T-LYMPHOCYTES SPECIFIC FOR EPSTEIN-BARR-VIRUS (EBV) NUCLEAR ANTIGENS FAIL TO LYSE THE EBV-TRANSFORMED B-LYMPHOBLASTOID CELL-LINES AGAINST WHICH THEY WERE RAISED

We have raised CD8(+) cytotoxic T lymphocytes (CTL) from three Epstein -Barr virus-seropositive donors by incubating peripheral blood lymphoc ytes with irradiated autologous B95.8-strain EBV-transformed B lymphob lastoid cells (LCL). However, to detect lysis in a standard Cr-51 rele ase assay of the LCL against which these CTL were raised, superinfecti on with recombinant vaccinia expressing the appropriate EBV protein or incubation with the peptide epitope was necessary. The untreated LCL were not lysed, even though Western blotting demonstrated that they ex pressed the EBV antigens containing the CTL epitopes. We have found CT L of this phenotype that are restricted by human leukocyte antigen-A2, -A3, -B7, or -B39, and which recognize the EBV latent proteins, EBV n uclear antigen (EBNA)-3A, EBNA-3C, or terminal protein. During these e xperiments, we identified a new human leukocyte antigen-A3-restricted EBNA-3A epitope, residues 603-611, RLRAEAGVK. We raised a spontaneous LCL, transformed by endogenous EBV, from one donor, but this was also not lysed. For at least one of the epitopes, CTL from another donor ly sed the LCL without superinfection or addition of peptides. We conclud e that the CTL were unable to achieve a high enough avidity of interac tion with untreated LCL to trigger effector function, although the LCL were able to stimulate them to grow in vitro for up to 4 mo. To asses s whether a small percentage of the LCL might possess a higher antigen density, we used an assay of tumor necrosis factor release from a CTL clone, which was able to detect antigen-bearing cells representing on ly 1% of a stimulating LCL population. Nevertheless, the untreated aut ologous LCL line failed to stimulate tumor necrosis factor release.