Figure 3

Activation of CD4+ T cells after primary and booster YF-17D vaccination. PBMCs were analysed by flow cytometry, and activated CD4⁺ T cells were defined as CD38⁺ HLA-DR⁺ CD4⁺ CD3⁺ T cells (see supplementary Figure S2, right-hand panel). (A) Frequencies of activated CD4⁺ T cells before and after YFV vaccination. Pre- (day 0) and post-vaccination (day 9 to 41) PBMCs from 209 primary YFV vaccinated donors were analysed. In the left-hand panel, pre-vaccination frequencies are plotted; in the right-hand panel, the post-vaccination frequencies are plotted according to the day the blood sample was collected. The median for each collection day is indicated. Mann Whitney U test was used to determine the significance of the difference between day 0 pre-vaccination, and the indicated day post vaccination (***p < 0.001; **p = 0.004; if nothing is given the sample size is too small). (B) The activation level of CD4⁺ T cells from primary and booster vaccinated donors before and after YFV vaccination. CD4⁺ T cell activation status was determined for PBMCs from primary vaccinated donors before (“naïve”; n = 147) and after (“primary effectors”; n = 178) vaccination and for PBMCs from booster vaccinated donors before (“late memory”; n = 29) and after (“secondary effectors”; n = 29) vaccination. The median is indicated. A Mann Whitney U test was used to determine the significance of the difference between the indicated groups (***p < 0.0001; **p = 0.001; *p = 0.0264; ns: not significant). (C) The total YFV specific CD4+ and CD8+ T cell responses in primary and booster vaccinated donors were analysed. PBMCs from primary vaccinated donors (“primary effector”; n = 100) and from booster-vaccinated donors (“secondary effector”; n = 18) were analysed ex vivo using IFN-γ specific ELISpot analysis using 30 peptide pools covering the entire YFV proteome. Only donors vaccinated 12–22 days prior to blood collection were included. For each donor, the total YFV specific T cell response was measured as the sum of IFN-γ specific cells (spot forming units, SFU per 10⁶ PBMC). The median is indicated. Mann Whitney U test was used to determine the significance of the difference between the groups (***p < 0.0001).