FIGURE 1.

Tfh cells are unique in promoting B cell differentiation and proliferation in vitro. (A) Sorting strategy for Tfh, Tfr or CXCR5- T reg cells. eGFP-Foxp3 mice were immunized with MOG/CFA and cells were obtained on day 7 post-immunization. Primed Tfh cells and Tfr cells were obtained from dLNs, sorted with CD4 beads, stained and sorted. Tfh cells were identified as CXCR5⁺ PD-1⁺ Foxp3^– CD4⁺ CD19^–, whereas Tfr cells were identified as CXCR5⁺ PD-1⁺ Foxp3⁺ CD4⁺ CD19^–. CXCR5-Tregs were identified as CXCR5^– PD-1⁺ Foxp3⁺ CD4⁺ CD19^– (arrow down). Foxp3 was identified by GFP (anti-CD25 and anti-CD44 was added when sorting for IL4-/- Tfh cells). (B,C) nvB were labeled with eF450 and cultured with primed Tfh or nvT cells and stimulated with anti-CD3 alone anti-CD3/anti-IgM. B cell proliferation was assayed by dilution of e450. (D) nvB cells alone or together with primed Tfh or nvT cells were stimulated with anti-CD3 alone or anti-CD3/anti-IgM. GC B cells were gated as live Fas⁺GL7⁺B220⁺ IgD^lo B220⁺IgD^–. (E) Switching was measured by staining GC B cells intracellularly with anti-IgG1. Summary of 10 independent experiments measuring IgG1⁺ GC B cells. Statistical results are from 10 independent experiments showing SEM (n = 10). *p < 0.05, ***p < 0.0001, unpaired Student t-test.