Figure 4.

ACAD8-T105I, BCAR1-G23V and PLCG1-M425Linduce more efficient NRT responses than WT epitopes in HLA-A2.1/K^bTg mice. (A–D) Splenocytes of mice(n=5) vaccinated with mutated peptides were tested by ELISPOT assay for the recognition of mutated peptides compared with the corresponding wild-type sequences. The data are presented as the means ± s.e.m.s from three independent experiments. **P < 0.01 was obtained in the comparison of IFN-γ (A, B), TNF-α (C) and IL-2 (D) production by splenocytes stimulated without a peptide or with VSV-NP43-69. (E–G) Splenocytes from HLA-A2.1/K^bTg mice immunized with mutated peptides were re-stimulated in vitro with the corresponding mutated peptide for 7 days. The ex vivo cytotoxicity against the corresponding mutated peptide-pulsed T2/Kb cells and minigene-nucleofected H522/Kb cells at the indicated E:T ratio was examined using time-resolved fluorescence assay. Irrelevant peptide VSV-NP43-69-pulsed T2/Kb cells, T2/Kb cells alone or H522/Kb cells were used as controls. Stimulation with anti-CD3-mAb was used as positive control. The data are presented as the means ± s.e.m.s from three independent experiments. **P<0.01. E:T, effector: target; SFC, spot-forming cell; VSV-NP, vesicular stomatitis virus nucleoprotein.