Dengmin Feng, Herbert Whinna, Dougald Monroe, and Darrel W. Stafford Abstract Activated factor VII is approved for treating hemophilia patients with autoantibodies to their factor IX or FVIII; however, its mechanism of action remains controversial. Some studies suggest that FVIIa requires tissue factor (TF) for function and that the reason for the high dose requirement is that it must compete with endogenous FVII for tissue factor. Others suggest that FVIIa binds platelets where it activates FX directly; the high concentration required would result from FVIIa’s weak affinity for phospholipids. We address this question by infusing a chimera of mouse FIX (Gla and EGF1) with FVIIa (EGF2 and catalytic domain) into hemophilia B mice. This mutant has no TF–dependent activity because it cannot functionally bind TF at physiologically relevant concentrations. In vivo, this mutant is as effective as mouse FVIIa in controlling bleeding in hemophilia B mice. Our results suggest that the hemostatic effect of pharmacologic doses of FVIIa is TF independent.