T-bet–dependent migration of CXCR3⁺ AFCs and GC B cells toward CXCL10. Chimeras were prepared by transfer of OTII cells alone or with OTI cells into congenic WT mice. Other chimeras were constructed by transfer of both OTII and OTI cells into T-bet^−/− mice. Seven days after immunization with alumOVA, suspensions of cells from the draining popliteal LN of the different chimeras were prepared. The chemotactic properties of the cells toward CXCL10 (a ligand for CXCR3) or CXCL12 (a ligand for CXCR4) were tested in migration assays using transwells containing 300 ng/mL of chemokine or medium in the bottom chamber. (A) The bar charts represent the percentage of AFCs (top chart) or GC B cells (bottom chart) expressing CXCR3 in the input cell suspensions (gray bars), or the mean ± SD (percentage) in the top chambers (black bars) and in the bottom chambers (white bars) obtained from 2 independent migration assays. The significance of differences was assessed by 2-tailed Mann-Whitney nonparametric statistics. NS indicates not significant. (B) The number of AFCs and GC cells found in the bottom chamber was determined by flow cytometry, and the migration index of the cells was calculated by reference to the number of cells found in the bottom chamber of the control with medium only. The bar charts represent the mean ± SD from triplicates in 1 representative experiment of 3. The figure shows 1 representative experiment of 3 independent experiments. The P values are given for statistically significant differences as calculated by t test.