Isotype switching and plasmablast differentiation in Igh^CGG/+ mice in vitro. (a) In vitro isotype switching. Purified Igh^CGG/+ (CD45.2/2) and WT (CD45.1/1) B cells were cocultured in vitro with LPS and IL-4 to induce isotype switching to IgG₁. Histogram shows IgG₁ staining by flow cytometry on day 3 of culture. Data are from three independent experiments and are quantified in the graph; proportions of IgG₁⁺ cells in Igh^CGG/+ and WT B cells from the same experiment are connected by a line. P = 0.74 (unpaired t test). (b) AFC cell formation (as defined by CD138 expression) and IgG₁ secretion in purified B cells stimulated with LPS and IL-4 in vitro. Cells were presorted as in Fig. 3 a, and non–CGG-binding B cells were excluded from Igh^CGG/+ cultures. Histograms are representative of, and graphs show pooled data for two independent experiments. n = 3 mice for WT and n = 6 mice for Igh^CGG/+ CGG binding B cells. P = 0.33 for CD138 expression and P = 0.37 for supernatant IgG₁. Numbers within flow plots indicate percentage of cells in the designated gate.