Expression of VWF-R1597W mutant and its cleavage by ADAMTS13 in the presence of mAb SZ34 under static/ nondenaturing conditions The plasmid PSVHvWF1, which harbors a full-length cDNA insert of human VWF (kindly provided by JE Sadler, Washington University School of Medicine, St Louis, USA) [29] , was used to construct the R1597W mutant by PCR-based single-nucleotide mutagenesis. Recombinant VWF-R1597W mutant was expressed in transiently transfected Hela cell line using serum-free OPTI-MEM I (Invitrogen) for 48 h. The media were collected and added into EDTA-free 16 proteinase inhibitor Cocktail (Roche), followed by concentration using Amicon Ultra-15 Centrifugal Filter (Millipore). VWF antigen (VWF:Ag) concentration was determined by an ELISA kit (Dako).