Supplementary Figure 1: The effects of BCR and or TLR9 signaling in B cell functions.

(a) Representative flow cytometry plots showing the differential phosphorylation of kinases, described in Fig. 1a–d. (b) The effects of Anti-IgM concentration on the phosphorylation of downstream kinases are shown. B cells were stimulated with 1 or 10 µg/ml Anti-IgM either individually or in combination with 1µM CpG. Phosphorylation of kinases was measured using phosphoflow as described in Fig. 1. Data are representative of two independent experiments performed in duplicates(a) or triplicates (b). (c) Changes in the phosphorylation of kinases in B cells obtained from TLR-9 KO mice in comparison with WT mice upon stimulation with CpG are shown. Data represent two independent experiments each carried out with duplicates (d) Calcium response of MD4 HEL transgenic B cells upon stimulation with 1 µg/ml HEL and/or 1 µM CpG are shown using a strategy described in Fig. 1e. Data are representative of three independent experiments. (e) Representative flow cytometry plots indicating the proliferation levels for B cells incubated with Anti-IgM (1 µg/ml) or CpG alone (0.04 µM) or with both from the experiment outlined in Fig. 1h. (f)The proliferative response of B cells in response to a range of Anti-IgM (Fab’2) antibody concentrations are shown. Data are representatives of two independent experiments each carried out with triplicates. (g) Percentage of IgG positive B cells in the live B cell population upon negative magnetic separation (left) or negative magnetic separation with additional IgG depletion (right) are shown. Data are representatives of two independent experiments.