Impaired human B-cell differentiation into Ig-secreting plasma cells by ectopic expression of Spi-B. CD19⁺ B cells isolated from peripheral blood were transduced with Spi-B, BCL-6, Spi-B∼ER, or control vectors and cultured in conditions promoting plasma cell differentiation (as in Figure 1). (A,B) After 7 days of culture, GFP⁺ cells were analyzed for CD19, CD20, CD38, and CD138 surface expression by flow cytometry. (A) Contour plots of one representative experiment of 10 are shown. Numbers in the quadrants indicate percentages of cells. (B) Percentages of CD38⁺CD20⁻ and CD19⁻CD138⁺ cells in Spi-B– and BLC-6–transduced cultures were normalized to control cultures. Mean plus or minus SD values of 10 independent experiments are shown (Student t test, ** P < .01). (C) Five days after transduction, GFP⁺ cells were sorted, and equal numbers of cells (50 000) were cultured with IL-2 and IL-21 for an additional 48 hours. The supernatants were collected, and IgM and IgG protein levels were analyzed by ELISA. Mean plus or minus SD values of ELISA triplicates are shown. One representative experiment of 3 is displayed. (D) After 7 days of culture, the percentages of CD38⁺CD20⁻ cells within the GFP⁺ population in Spi-B∼ER and control transduced cultures supplemented with the indicated concentrations of 4HT were assessed. One representative experiment of 2 is shown.