Von Willebrand factor (VWF) is an essential contributor to microvascular thrombosis. Physiological cleavage by ADAMTS13 limits its prothrombotic properties, explaining why ADAMTS13 deficiency leads to attacks of microthrombosis in patients with thrombotic thrombocytopenic purpura (TTP). We previously reported that plasminogen activation takes place during TTP attacks in these patients. Furthermore, stimulation of plasminogen activation attenuates pathogenesis in preclinical TTP models in vivo. This suggests that plasmin is an endogenous regulator of VWF thrombogenicity, in particular when ADAMTS13 falls short to prevent microvascular occlusions. VWF cleavage by plasmin is biochemically distinct from cleavage by ADAMTS13. We hypothesized that plasmin-cleaved VWF (cVWF) holds value as a biomarker of microvascular thrombosis. We here describe the development of a VHH-based bioassay that can distinguish cVWF from intact and ADAMTS13-cleaved VWF in plasma. We validate this assay by tracking cVWF release during degradation of microthombi in vitro. We demonstrate that endogenous cVWF formation takes place in TTP patients during acute attacks of thrombotic microangiopathy, but not in remission. Finally, we show that therapeutic plasminogen activation in a mouse model for TTP amplifies cVWF formation, which is accompanied by VWF clearance. Our combined findings indicate that cVWF is released from microthrombi in the context of microvascular occlusion.