B cells express GITR starting at the transitional stage of development.

<p>(A) Bone marrow and spleen cells from wild-type (129S1) mice were analyzed <i>ex vivo</i> by flow cytometry for the expression of indicated markers. Live cells in dot plots were gated as indicated on top. Histograms depict GITR expression on B cells in subsets gated as shown in the dot plots in top panel. Expression of GITR on gated CD4⁺ T cells is shown as a positive control for GITR staining. Rat IgG2b was used as an isotype control for GITR staining. Data are representative of 3 independent experiments. Similar GITR expression was observed on B cells from BALB/c mice (data not shown). (B) GITR^+/+ (BALB/c) mice were immunized i.p. with 100 µg NP₃₉-CGG in Alum. At days 4, 7, and 15 post immunization, spleen cells were harvested from 2 immunized and 2 nonimmunized (naïve) mice at each point and analyzed for GITR expression on germinal center B cells reactive with NP. The dot plots show NP and PNA binding on B220⁺PNA^high gated B cells from one naive (left panel) and one immunized (right panel) mouse at day 7. There were no NP⁺ germinal center B cells at day 4, and reduced frequency at day 15 (data not shown). The histogram represents live, B220⁺IgD⁻CD3⁻CD11b⁻PNA^high germinal center B cells. GITR expression on NP⁺ germinal center B cells of one mouse at day 7 (black intact line) and one mouse at day 15 (gray intact line) following immunization are shown compared to those on total B220⁺PNA⁻ B cells (dark-shaded histogram and dashed black line) in respective animals and on lymphocytes of one GITR^−/− mouse (light-shaded histogram).