PPD responders and BCG vaccinated infants recognize predicted HLA-E binding peptides derived from Mtb.

<p>(A) Summary of all proliferation data from 10 PPD responders and 10 PPD non-responding donors. Proliferation was calculated using the following formula: Geometric mean of undivided population minus geometric mean of all cells. Subsequently the percentage was calculated by: (delta geo mean of sample- delta geo mean of negative control)/ delta geo mean of maximal proliferation (PHA). Samples with 10% specific proliferation or more were considered positive (black boxes). Blue highlights identify the peptides that were selected for detailed analysis. Umbilical cord blood (UCB) (n = 5) cell proliferation was assessed by the percentage of proliferating CD3⁺CD8⁺CD56⁻ cells and proliferation >10% was considered positive. (B) PBMCs from BCG vaccinated infants (n = 12) were tested in the same way as the cells from PPD responders for 10 selected peptides. Proliferation of 5–10% is indicated in grey and proliferation >10% in black. (C) Comparison of PPD responders, non-responders & cord blood samples for peptide recognition, i.e. the number of peptides recognized per donor (n = 10 for adults, n = 5 for cord blood). Analyzed using Students T-test with a p<0.05 considered significant. (D) CD8⁺ T cells were purified from PBMCs by magnetic bead separation of donors 2,4 and 6 and proliferation (CFSE) in response to peptide pre-pulsed K562 cells with (right plots) or without (left plots) HLA-E was assessed. Plots are gated on CD8⁺CD56⁻ cells and CFSE dilution is plotted.