Comparison of PR1 and PR2 peptides for HLA-A2–binding affinity and stability of the MHC-I complexes on the surface of T2 cells. (A) Flow cytometric histogram showing labeling of HLA-A2 on the surface of T2 assays using FITC-conjugated Ab to HLA-A2 following incubation of the cells with 100μM PR1 or PR2 peptides. A known HLA-A2 binding CMV pp65 CTL epitope peptide and a HLA-B7–restricted CMV peptide were used as positive and negative controls. (B) Titration of peptide concentration in the assay of panel A. The binding affinity of the pp65, PR1, and PR2 epitope peptides for HLA-A2 was not significantly different as determined by 1-way ANOVA tests. (C) Time-course experiment to investigate stability of peptide-HLA-A2 complexes following washout of peptide from the T2 cell cultures. Experiments were performed twice each, at peptide concentrations of 3, 6, 12, 25, and 50μM. Panel C shows the experiment using 25μM peptide concentration. The other conditions gave comparable data (data not shown). For further details, see “Peptide-binding assay using T2 cells.”