Evaluation and clinical application of a new method for measuring activity of von Willebrand factor-cleaving metalloprotease (ADAMTS13)

Thrombotic thrombocytopenic purpura (TTP) is associated with acquired or congenital deficiency of a plasma von Willebrand factor-cleaving protease (VWFcp). Based on partial amino acid sequence and genomewide linkage analysis of pedigrees with congenital TTP, VWFcp was recently identified as a new member of the ADAMTS family and designated ADAMTS13. We developed a new, rapid, and simple method for measuring VWFcp activity based on the positive correlation between VWF multimeric size and Ristocetin cofactor activity (VWF:RCo). After dilution of plasma with low ionic Tris buffer and activation of the protease with barium chloride, a VWF concentrate is digested in the presence of urea. Subsequently, the residual VWF:RCo of the samples is assessed and used to calculate the VWFcp activity of the samples. The accuracy of the new technique is verified by estimating VWFcp activity for 282 plasma samples with the RCo-based assay and the original immunoblotting assay. The method is reproducible as shown by low intra-and interassay coefficients of variation (2.8% and 7.5% for normal samples, respectively, and 8.7% and 12.9% for abnormal samples, respectively). Furthermore, the clinical application of the new method is illustrated by measuring VWFcp of 14 patients with 22 episodes of acute TTP as well as other thrombotic, thrombocytopenic, or hemolytic disorders. Severe VWFcp deficiency was restricted to patients with acute, classic TTP. The majority of patients with low titer inhibitor respond to plasma exchange treatment with increase of VWFcp activity, whereas VWFcp deficiency persists in some patients with high titer inhibitor despite clinical remission.