American Society of Hematology

Figure 2

$Figure 2. CD40 induced NFκB activation. (A) CD19+ splenic B cells purified from different CD40 transgenic mice were cultured in vitro with hCD40L (1 μg/mL) stimulation for indicated time points. Canonical NFκB activation was measured by phosphorylated IκBα blotting. Anti-α/β actin antibody was used for loading control. (B) For noncanonical NFκB activation, B cells were cultured in 24-well plates and stimulated with 1 μg/mL hCD40L for 16 hours. Cells were then harvested and cytoplasmic and nuclear fractions were isolated. The level of noncanonical NFκB activation was detected by p52 blotting in the nuclear fraction. Anti-SAM68 antibody was used for loading control. Data are representative of more than 3 independent experiments. (C) The ratio of intensity values of p52 and SAM68 signals from the nucleus fraction was presented as quantitative analysis for measuring noncanonical NFκB activation.$

CD40 induced NFκB activation. (A) CD19⁺ splenic B cells purified from different CD40 transgenic mice were cultured in vitro with hCD40L (1 μg/mL) stimulation for indicated time points. Canonical NFκB activation was measured by phosphorylated IκBα blotting. Anti-α/β actin antibody was used for loading control. (B) For noncanonical NFκB activation, B cells were cultured in 24-well plates and stimulated with 1 μg/mL hCD40L for 16 hours. Cells were then harvested and cytoplasmic and nuclear fractions were isolated. The level of noncanonical NFκB activation was detected by p52 blotting in the nuclear fraction. Anti-SAM68 antibody was used for loading control. Data are representative of more than 3 independent experiments. (C) The ratio of intensity values of p52 and SAM68 signals from the nucleus fraction was presented as quantitative analysis for measuring noncanonical NFκB activation.

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