B cells stimulated with F(ab′)2 or WM anti-IgM show similar levels of activation and lack of BLyS receptor up-regulation is not because of increased cell death. (A) CD23+ B cells were cultured with the indicated stimuli (concentrations of stimuli are indicated in “B-cell subset isolation, culture, and stimulation”). After 24 hours, B cells were harvested and analyzed by FACS for forward side scatter/side scatter (FSC/SSC), CD86, and CD69 expression on live cells. Blasting profiles (far left column) have FSC displayed on the x-axis and SSC displayed on the y-axis. Live cells were gated and the percentage of live cells shown. Histograms of CD86 and CD69 expression on live cells are shown in the middle and right columns. (B) CD23+ B cells from Bcl-xL transgenic or control C57BL/6 mice were cultured with the indicated stimuli. After 24 hours, the cells were harvested and analyzed by FACS for BR3 and TACI expression on live cells. Gray histograms indicate isotype control; unfilled white histograms, levels expressed by unstimulated; filled black histograms, levels expressed by stimulated cells. Median fluorescence intensities (MFI) of respective cell populations are shown in bar graphs at bottom. Plots are representative of 3 independent experiments; in all cases, n equals 3 or more. Significance levels are indicated by P values.
