Purified CD11c⁺ DCs in the presence of TLR7 agonist facilitate T cell–independent restimulation of FVIII-specific memory B cells. Memory B cells were obtained from hemophilic mice treated with 4 weekly doses of FVIII. DCs and naive B cells were obtained from untreated hemophilic mice. (A) Purified memory B cells and purified CD11c⁺ DCs were cultured in the presence of FVIII only, FVIII and TLR7 agonist imiquimod (100 ng/mL), or FVIII and a mixture of TLR7 agonist imiquimod (100 ng/mL) and TLR9 agonist CpG-ODN (100 ng/mL). The concentration of FVIII is indicated. Newly differentiated FVIII-specific ASCs were analyzed after 6 days of culture. Representative ELISPOTs are presented. (B) Purified naive B cells and purified CD11c⁺ DCs were cultured in the presence of FVIII only, FVIII and TLR7 agonist imiquimod (100 ng/mL), or FVIII and a mixture of TLR7 agonist imiquimod (100 ng/mL) and TLR9 agonist CpG-ODN (100 ng/mL). The concentration of FVIII was 10 ng/mL. Newly differentiated FVIII-specific IgG ASCs and IgM ASCs were analyzed after 6 days of culture. Representative ELISPOTs are presented.