Mapping of dominant hF.IX CD4⁺ T-cell epitopes. (A) Representative result of T-cell proliferation following in vitro stimulation with individual peptides of a hF.IX peptide library. Splenocytes were isolated from hemophilia B C3H/HeJ mice after immunization with hF.IX in cFA. Incorporation of ³H thymidine was measured and compared with mock- and hF.IX protein–stimulated cells. cpm indicates counts per minute. (B) Diagram of hF.IX primary amino acid sequence. Boxes indicate peptides that contain dominant CD4⁺ T-cell epitopes in hemophilia B mice on C57BL/6, C3H/HeJ (this study), or BALB/c (Greenwood et al²⁵ ) genetic background. Also indicated are domains and protease cleavage sites and numbering of some reference amino acids.