Supplementary Figure 1: Characterization of the T_FR-DTR strain.

a) Gating strategy to identify T_FH, T_FR and CXCR5^– T_reg cells in draining lymph nodes of mice immunized with NP-OVA 7 days previously. b) DTR expression on T_FH, T_FR and CXCR5- T_reg cells gated as in (a) in indicated mouse strains. Full stain = all antibodies used along with secondary reagent for DTR staining. No DTR = all antibodies except anti-DTR primary was not added. c) Quantification of activated T_reg cells in T_FR-DTR mice. Control or T_FR-DTR mice were immunized and given DT to delete T_FR cells as in Fig. 1f. CXCR5-Ki67⁺FoxP3⁺ activated T_reg cells were quantified. Representative gating (left) and analysis (right) are shown. d) Quantification of T_FR cells by flow cytometry (left) and quantification of FoxP3⁺ cells within individual GCs by microscopy (right) in T_FR-DTR mice as in (c). Column graphs represent the mean with error bars indicating standard error. P value indicates two-tailed student’s T test. Data are from an individual experiment and are representative of two experimental repeats (a-c), or are combined data from two experiments (d).