Figure 5
Figure 5. In vitro CSR analysis in ArteΔ/− B cells. Surface expression of switched isotypes. Purified B cells from Artfl/− and ArtΔ/− mice were labeled with CFSE and stimulated for 4 days with various polyclonal B-cell activators for switching to IgG1, IgE, IgG2b, IgG3, and IgA (“Purification and activation of splenic mature B cells in vitro”), and stained with anti-B220 and specific anti-Ig antibodies for flow cytometric analysis. All cells were B220+, assessing for the efficiency of B-cell purification. (A) Percentage of total B cells expressing IgG1, IgE, IgG2b, IgG3, and IgA, as determined by flow cytometric analysis on B cells from Artfl/− (▴) and ArtΔ/− mice (▵), after 4 days of in vitro stimulation. **Statistically significant difference (P > .003, 2-tailed Mann-Whitney test). (B) Analysis of CSR in the various subsets of proliferative B cells according to CFSE intensity. Plots show IgG1, IgE, IgG2b, IgG3, and IgA staining together with CFSE, on viable (propidium iodide–negative) lymphoid cells. The percentages of switched cells are indicated. (C-D) Percentages of total (C) and IgG1-, IgE-, IgG2b-, IgG3-, or IgA-switched (D) B cells that have undergone the indicated numbers of cell division, for Artfl/− and ArtΔ/− mice. Columns and bars represent mean and SD of independent animals depicted in panel A.

In vitro CSR analysis in ArteΔ/− B cells. Surface expression of switched isotypes. Purified B cells from Artfl/− and ArtΔ/− mice were labeled with CFSE and stimulated for 4 days with various polyclonal B-cell activators for switching to IgG1, IgE, IgG2b, IgG3, and IgA (“Purification and activation of splenic mature B cells in vitro”), and stained with anti-B220 and specific anti-Ig antibodies for flow cytometric analysis. All cells were B220+, assessing for the efficiency of B-cell purification. (A) Percentage of total B cells expressing IgG1, IgE, IgG2b, IgG3, and IgA, as determined by flow cytometric analysis on B cells from Artfl/− (▴) and ArtΔ/− mice (▵), after 4 days of in vitro stimulation. **Statistically significant difference (P > .003, 2-tailed Mann-Whitney test). (B) Analysis of CSR in the various subsets of proliferative B cells according to CFSE intensity. Plots show IgG1, IgE, IgG2b, IgG3, and IgA staining together with CFSE, on viable (propidium iodide–negative) lymphoid cells. The percentages of switched cells are indicated. (C-D) Percentages of total (C) and IgG1-, IgE-, IgG2b-, IgG3-, or IgA-switched (D) B cells that have undergone the indicated numbers of cell division, for Artfl/− and ArtΔ/− mice. Columns and bars represent mean and SD of independent animals depicted in panel A.

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