IgG1 anti-CD20 treatment plus high-dose FVIII exposure up-regulated B7.2 and B7.1 costimulatory molecules on the MZ B cells. Groups of FVIII primed E16 mice (n = 9-11) were treated with either IgG1 anti-CD20 or control IgG (250 μg per mouse), followed by twice daily intravenous injection of 2 μg FVIII for 5 days. One week after the final intravenous FVIII injection, spleen B cells were analyzed for B7 molecule expression. A group of naive mice (n = 8) was included as an additional control. (A) Representative dot plot shows the gates for FO (CD23^hiCD21^low) and MZ (CD23^intCD21^hi) B cells. Cells were gated on live CD19⁺ B cells. (B) Representative overlay histographs show the B7.2 and B7.1 expression on MZ B cells from naive mice (solid gray), mice with control IgG pretreatment (line), and mice with IgG1 anti-CD20 pretreatment (dashed line). (C) Bar graphs represent the quantitative results of mean fluorescence intensity (MFI) for B7.2 and B7.1, respectively. *P < .05 (Student t test, 2-tailed). n.s. indicates not significant.