Differential expression of IL-17F and IL-17A by Vγ6/Vδ1⁺ γδ T cells. (A) IL-17F concentration in lung homogenates was analyzed by ELISA from WT C57BL/6 (wt), B6.Vγ6^+/+ (+/+), and B6.TCRδ^−/− (δ^−/−) mice treated with either B. subtilis or PBS for 4 wk. The mean is shown as a solid line. Data are compiled from five individual mice from at least two separate experiments. (B) Representative density plots of intracellular IL-17F and IL-17A expression in CD4⁺ T cells isolated from the lung of WT C57BL/6, B6.Vγ6^+/+, and B6.TCRδ^−/− mice treated with B. subtilis for 4 wk. CD4⁺ T cells that express IL-17F and IL-17A were selected from a bivariate dot plot of CD3⁺/CD4⁺-expressing cells in the lymphocyte gate. The percentage of CD4⁺ T cells expressing IL-17A or IL-17F is shown in each quadrant of the density plot. (C) Absolute number of IL-17A– or IL-17F–expressing CD4⁺ T cells in the lung of WT C57BL/6, B6.Vγ6^+/+, and B6.TCRδ^−/− mice treated with B. subtilis for 4 wk. Data represent at least five individual mice per experiment from at least three separate experiments (mean ± SD). (D) Representative density plots of intracellular IL-17F and IL-17A expression in γδ T cells isolated from the lung of WT C57BL/6 and B6.Vγ6^+/+ mice treated with B. subtilis for 4 wk. γδ T cells expressing IL-17A, IL-17F, or IL-22 were selected from a bivariate dot plot of CD3⁺/Cδ⁺ cells from the lymphocyte gate. The percentage of γδ T cells expressing IL-17A, IL-17F, or IL-22 is shown in each quadrant of the density plot. (E) Absolute number of IL-17F–expressing γδ T cells in the lung of B6.Vγ6^+/+ and WT C57BL/6 treated with B. subtilis for 4 wk using different gating strategies labeled A, B, or C. Data represent at least five individual mice per experiment from at least three separate experiments (mean ± SD).