CTL line A2-AL9 specifically killed activated primary CD4+ T cells (expressing HLA-A*02:01 and HLA-B*35:01 and used as antigen presenting cells) loaded with cognate peptide AL9 but not non-cognate peptide SL9 (A). Peptides that binds specifically to HLA-B*35:01 (YPL, HPN, HPV and FPT) could reduce AL9-induced cell lysis. Representative data with CD4+ T cells from Donor 24 are shown in Panel A and statistical analyses from Donors 24 and 168 in Panels B and C. Cell lysis assays were also performed with another CTL line B8-RL8 and activated primary CD4+ T cells (carrying HLA-B*08:01 and B*35:01) from Donors 25 and 132 loaded with cognate peptide RL8 were used as target cells (D and E). (B–E) The mean ± SEM of three independent assays are shown. Statistical analyses were undertaken using one-way ANOVA analysis with Dunnett test. *, p<0.05, **, p<0.01, ***, p<0.001, ****, p<0.0001. (F–I) Blocking of HLA-B*35:01 peptide-deficient conformers on the surface of CD4+ T cells by B*35:01-specific peptides (YPL, HPN, HPV and FPT), but not nonspecific control peptide (TW10 or HGV) was confirmed by flow cytometry after staining cells with HC10. The mean ± SEM of two to three independent experiments are shown.