Human GC B-cell purification (A.1), annexin-V binding expression kinetics (A.2), and surface molecule expression (B). (A.1) Unfractionated and GC B cells were obtained from human tonsils and the expression of CD20 and CD38 on their surface was monitored by immunofluorescence and flow cytometry. Dot plots corresponding to a representative experiment are shown. (A.2) GC B cells (10⁶cells/mL) were cultured for indicated times in the absence and in the presence of anti-CD40 MoAb (1 μg/mL) + IL-4 (2 ng/mL) or VAD-fmk (200 μmol/L) and the cells were then labeled with FITC-annexin-V and analyzed by flow cytometry. Histograms of annexin-V expression in one representative experiment are shown. (B.1) GC B cells were cultured for 2 hours and simultaneously labeled with FITC-annexin-V and additional MoAb directed to the indicated molecules. Dot plots of one representative experiment are shown. All of the surface molecules examined were positive in more than 90% of the freshly isolated GC B cells, except for CD49d, which was positive in only 39% ± 5% (mean ± SEM) of the cells. Axis scales of dot plots are logarithmic. (B.2) Expression of surface molecules in annexin-V⁺ GC B cells. The values were obtained as the percentage of the MFI shown for each surface molecule studied on the annexin-V⁺ cells, with respect to that observed on annexin-V⁻ cells, which was considered the control expression. Results represent the mean ± SEM of five experiments.