Fig. 4

Serum-free and stromal cell-free culture of CLP cells ex vivo. (A) Microscopic observation of B lymphopoiesis of CLP in vitro. 1,000 sorted CLP were cultured in stromal free and serum free conditions with 20 ng/ml IL-7、100ng/mL FLT3L and 40ng/mL SCF for the indicated times; a, b, c, d demonstrated the microscopic manifestations of CLP differentiation into B cells on day 0, day 3, day 5 and day 7, respectively; a, b scale bars indicate 150 μm; c, d scale bars indicate 750 μm. (B) Diagram of B cell flow gating strategy. (C) In vitro B lymphopoiesis of CLP in the presence of IL-7, FLT3L and SCF. The development of B-lineage cells was analyzed via flow cytometry with antibodies against B220, CD19, CD43 and IgM during cultured for 7 d. (D) The increase in the number of total cell, CD19 + B220+, CD19 + B220 + IgM + cells and each cell population under culture. The percentage mean each cell accounted for the total cell frequency. *P<0.05. **P<0.01. ***P<0.001. ****P<0.0001