ADAMTS13 is a protease that cleaves the von Willebrand Factor (VWF) within intact blood vessels under shear stress 
. VWF is a large glycoprotein secreted by vascular endothelial cells as multimers. At a region of vascular injury, the multimeric form of VWF initiates the clotting process by adhering to platelets. A reduction or elimination of the protease activity of ADAMTS13 results in the VWF multimers remaining uncleaved in the circulating blood stream, which ultimately leads to intravascular thrombosis and an associated disorder known as Thrombotic Thrombocytopenic Purpura (TTP) 
. Thus ADAMTS13 plays a critical role in maintaining intravascular homeostasis.
ADAMTS13 is a member of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family of proteins 
and is secreted by almost all tissues but primarily by hepatic stellate cells 
. The ADAMTS family of proteins are secreted metalloproteases characterized by multiple domains 
. The domain structure of ADAMTS13 contains a signal peptide, propeptide, a metalloprotease, disintegrin, a thrombospondin type 1 domain (TSP1), a cystein rich domain, a spacer domain, seven TSP1 repeats and two C-terminal CUB (C1r/C1s, Urinary EGFand Bone morphogenetic protein) domains. Although all ADAMTS proteins have characteristic multi domains, the ADAMTS13 is unique in having two additional C-Terminal CUB domains and an unusually short propeptide. Moreover, unlike other ADAMTS members, ADAMTS13 is catalytically active prior to secretion from the cells 
. The metalloprotease domain is the catalytic domain and responsible for the protease activity 
. The region from the disintegrin to the spacer domains is involved in substrate recognition 
while the distal C-terminal TSP1 repeats and CUB domains are also necessary for its activity under flowing conditions 
. The role of the cysteine-rich domain is more controversial 
. It is thus evident that the ADAMTS13-VWF interaction is complex; several domains of the protein are involved in the regulation of VWF cleavage.
Antibodies sensitive to the conformation of a protein have been successfully used to understand the structural organization of proteins, distinguish their functional and non-functional forms, elucidate molecular mechanisms, and establish the role of different domains of multidomain proteins. For example, conformation-sensitive antibodies against native and denatured bovine somatotropin have been used successfully to study its folding, stability, thermal denaturation and refolding 
. The specific monoclonal antibody 5D3 was used to differentiate functional and non-functional ABCG2 proteins which had functional ATP- and drug-substrate-binding sites but differed in the formation of a catalytic intermediate 
. A monoclonal antibody against the CRIB domain of the N-WASP protein that specifically recognized the activated protein was used to localize it within cells 
. Conformation-sensitive UIC2 could identify different classes of drug modulators of P-glycoprotein and the molecular mechanism underlying their interactions based on mutations in the multidrug resistance gene (MDR
1) which led to different conformations of the molecule 
. Similarly, a conformation-sensitive antibody was successfully used to identify the disease-causing mutation in Copper/Zinc Superoxide Dismutase 
ADAMTS13 is a multi-domain protein with complex interactions with its substrate. Antibodies sensitive to different conformations could thus be potentially useful to understand the regulation and mechanism of ADAMTS13-mediated catalysis. In this report, we studied and defined new conformation-sensitive antibodies to ADAMTS13. We examined the monoclonal antibodies Wh2-11-1 and Wh2-22-1A, which are specific to the TSP1-4 and Disintegrin domains, respectively, and the polyclonal antibody BL154G that is specific to the metalloprotease domain of ADAMTS13, for their sensitivity to conformation. Here we show their ability to detect conformation changes in the ADAMTS13 protein under various conditions in which the protein normally undergoes changes in conformation such as changes due to temperature, and in the presence of its substrate. We use flow cytometry to measure the affinities and reactivity of these antibodies. We demonstrate that both the reactivity and the apparent affinity (concentration required for half-maximal binding) of these antibodies is altered by the addition of substrate (VWF). This change is observed mainly at 37°C but not at 4°C and occurs in the wild-type protein but not in a catalytically inactive mutant. These results indicate that the ADAMTS13 antibodies are conformation-sensitive.