Normal GC numbers, morphology, and cell cycle in SFR KO mice. Related to Fig. 2. (A–D) WT (n = 4) and SFR KO (n = 4) mice expressing AID-GFP were immunized i.p. with SRBCs. On day 7, spleen was removed, and cryosections were stained with either anti-IgD (B cells; red) plus anti-CD4 (CD4⁺ T cells; blue) antibodies (C) or biotin-anti-CD35 antibodies plus streptavidin-PE (red; D). Cells were analyzed by confocal microscopy. GC B cells were identified by detecting GFP (green). The protocol is depicted in A, and GC numbers per longitudinal section are shown in B. Representative photomicrographs of one experiment are provided in C and D. (E) The proportions of T reg cells (FoxP3⁺CD4⁺), Tfr cells (FoxP3⁺CD4⁺CXCR5⁺PD-1⁺), and GC Tfr cells (GL-7^hiFoxP3⁺CD4⁺CXCR5⁺PD-1⁺) in indicated mice were analyzed by flow cytometry (WT, n = 7; SFR KO, n = 6; SAP KO, n = 6; SFR-SAP dKO, n = 6), as for Fig. 4, A–D. Data are pooled from three independent experiments. (F) Mice were immunized with NP-OVA and, on day 7, cell cycle was analyzed by staining cells for Hoechst, which detects DNA. Percentages of GC B cells showing DNA >1n (cells in S-G2-M phases) are enumerated (WT, n = 6; SFR KO, n = 8). Data are pooled from two independent experiments. Each symbol represents one mouse. *, P < 0.05; **, P < 0.01 (two-tailed Student’s t test). Data are mean ± SEM. Scale bar, 50 µM. D, day; ns, not significant.