T cell–independent restimulation of FVIII-specific memory B cells requires cell–cell interactions between memory B cells and CD11c⁺ DCs. Purified memory B cells were obtained from hemophilic mice treated with 4 weekly doses of FVIII. CD11c⁺ DCs were obtained from untreated hemophilic mice. (A) Purified memory B cells only were seeded in the lower compartment. (B) Purified memory B cells and CD11c⁺ DCs were seeded in the upper compartment. Both compartments were restimulated with FVIII (10 ng/mL) together with TLR7 agonist imiquimod (100 ng/mL). Newly differentiated FVIII-specific ASCs were analyzed after 6 days of culture. Representative ELISPOTs are presented, each represents a pool of 4 wells.