Identification of immunogenic CCNA1 epitopes and HLA-restriction profiling studies. (A) Donor 2’s cell line was tested against each of the 21 minipools by IFN-γ ELISpot (SFC/1 × 10⁵). (B) Arrangement of peptides into minipools; donor 2’s reactive minipools (intersecting at peptides 45, 47, and 48) are shaded. Individual testing of these peptides identified peptide 48 as the stimulatory epitope (C), which was then confirmed by IFN-γ ICS (D). (E) Donor 2’s cell line lysed peptide-pulsed autologous and allogeneic targets matched at HLA-B7 but not at other alleles in a 4-hour Cr⁵¹ cytotoxicity assay, thus determining that peptide 48 is presented in context of HLA-B7. Results are presented as percent specific lysis, E:T 40:1.