The titers of anti-FVIII antibodies in hemophilia A mice after platelet depletion followed by rhF8 immunizations. (A) Schematic diagram of platelet depletion and rhF8 infusion in F8^null(B6/129S) mice. Ab, anti-GPIb antibody. Animals were infused IV with a single dose (2 mg/kg) of anti-GPIb antibody followed by 4 doses of rhF8 immunization (50 U/kg per week by IV injection). Isotype IgG was used in parallel as a control. (B) The inhibitor titers in F8^null(B6/129S) mice. One week after the last immunization, anti-FVIII inhibitors were determined by Bethesda assay. (C) The total anti-FVIII antibody titers in F8^null(B6/129S) mice. One week after the last immunization, total anti-FVIII antibody titers were determined by ELISA. (D) Schematic diagram of platelet depletion and rhF8 infusion in F8^null(B6) mice. Anti-GPIb antibody was infused at the first week at a dose of 2 mg/kg and the third week at a quarter dose (0.5 mg/kg) followed by rhF8 immunizations (50 U/kg per week IV ×4). Isotype IgG was used in parallel as a control. (Ei) The inhibitor titers in F8^null(B6) mice. One week after the last immunization, inhibitor titers were determined by Bethesda assay. (Eii) The titers of inhibitory antibodies in immunized F8^null(B6) mice with inhibitors ≤50 BU/mL. This is a zoom-in look at the lower part of panel Ei. (F) The total anti-FVIII antibody titers in F8^null(B6) mice. One week after the last immunization, total anti-FVIII antibody titers were determined by ELISA.