Fig. 7. ZnT8_186–194-reactive CD8⁺ T cells cross-recognize a B. stercoris mimotope.

(A–C) Four donors with sizable ZnT8_186–194 MMr⁺CD8⁺ T-cell fractions were selected. A first PBMC aliquot received PE/BV786-labeled ZnT8_186–194 MMrs and PE/BV711-labeled EboV NP_202–210 MMrs. For the second aliquot, PE-labeled B. stercoris MMrs replaced the PE-labeled ZnT8_186–194 MMrs. (A) Overlay of ZnT8_186–194/ZnT8_186–194 MMr⁺ (blue) and ZnT8_186–194/B. stercoris MMr⁺ cells (red). (B) Negative control staining of ZnT8_186–194/EboV NP_202–210 MMr⁺ cells. (C) Positive control staining of EboV NP_202–210/EboV NP_202–210 MMr⁺ cells from the first and second aliquot. The frequencies of MMr⁺ out of total CD8⁺ T cells are indicated. (D) Four ZnT8_186–194-reactive CD8⁺ T-cell clones (D222D 2, D349D 178B9, H017N A1, H328C 9C8) were stained with BV786/PE-labeled ZnT8_186–194, PE-labeled B. stercoris and BV650-labeled MelanA_26–35 MMrs. The ZnT8_186–194/B. stercoris cross-reactive clone H017N is shown, from left to right: ZnT8_186–194/B. stercoris MMr⁺; ZnT8_186–194/ZnT8_186–194 MMr⁺; and negative control ZnT8_186–194/MelanA_26–35 and B. stercoris/MelanA_26–35 MMr⁺ cells. (E) The H017N clone was stimulated with peptide-pulsed LCLs (0.1 µM, 6 h). Percent cytokine⁺ cells are shown as mean±SEM of two experiments. p=0.008 for Wilcoxon signed-rank comparison of pooled cytokine responses between B. stercoris and ZnT8_186–194, MelanA_26–35 or no peptide, and between ZnT8_186–194 and MelanA_26–35 or no peptide.