Recent data indicate an important contribution of coagulation factor (F)XII to in vivo thrombus formation. Since fibrin structure plays a key role in clot stability and thrombosis we hypothesized that FXII(a) interacts with fibrin(ogen) and thereby regulates clot structure and function. In plasma and purified system, we observed a dose-dependent increase in fibrin fiber density and decrease in turbidity, reflecting a denser structure, and a non-linear increase in clot stiffness with FXIIa. In plasma, this increase was partly independent of thrombin generation, as shown in clots made in prothrombin deficient plasma initiated with snake venom enzyme and in clots made from plasma deficient in FXII and prothrombin. Purified FXII and a-FXIIa, but not ß-FXIIa, bound to purified fibrinogen and fibrin with nanomolar affinity. Immunostaining of human carotid artery thrombi showed that FXII co-localized with areas of dense fibrin deposition, providing evidence for the in vivo modulation of fibrin structure by FXIIa. These data demonstrate that FXIIa modulates fibrin clot structure independently of thrombin generation through direct binding of the N-terminus of FXIIa to fibrin(ogen). Modification of fibrin structure by FXIIa represents a novel physiological role for the contact pathway which may contribute to the pathophysiology of thrombosis.