PMC

Proposed VWD diagnostic algorithm incorporating H5VWM. *The more of these features the stronger the recommendation for H5VWM and vice versa. ^†Follow orange line for VWF ≥50 IU/dL in absence of the mentioned features. AHA, acquired hemophilia A; FVIII, factor VIII activity; H5VWM, Hydragel 5 von Willebrand Multimer; MPS, myeloproliferative syndrome; PFA, platelet function analyzer; RIPA, Ristocetin‐Induced Platelet Aggregation; VWF, von Willebrand factor; VWD, von Willebrand disease; VWF ratio, ratio of VWF Act/VWF:Ag

(A‐E) Example Hydragel 5 VW multimers (H5VWM) densitometry of patients from the study. (A) Patient 10, normal VWF and 29% high molecular weight multimers (HMWM). (B) Patient 119 normal VWF, 22% HMWM and p.P1337L mutation linked to type 2B VWD. (C) Patient 70, compound heterozygous for p.F1501S and p.P1266E mutations but categorised as type 1 VWF with loss of IHVWM and normal H5VWM. (D) Patient 125 categorised as type 2M VWD but flattened HMWM peak and p.R1374C mutation. (E) an example of ultra‐large MWM in P165 with TTP and reduced ADAMTS13

Rationalization of patient results according to the VWF activity, VWF Ac, ratio of VWF Ac/VWF:antigen (VWF:Ag) and in‐house multimer, IHVWM, result. Patients were group according to VWF Ac and then ratio of VWF activity and VWF:Ag. IHVWM pattern was assessed as normal or reduced high molecular weight multimers (HMWM) when less than 11 multimer triplets were visible. The number of patients with discrepancy between IHVWM and Hydragel 5 multimer method, H5VWM, is indicated. Results from patients with type 2N, type 3 and acquired von Willebrand syndrome (AVWS) were excluded

(A‐D) Limit of detection of Hydragel 5 VW Multimers (H5VWM) using diluted normal plasma. H5VWM densitometry from left low molecular weight multimers (LMWM) to right high molecular weight multimers (HMWM) with peaks 1‐3 LMWM, 4‐7 intermediate molecular weight multimers (IMWM), and above 7 HMWM. Normal quality control sample is indicated by a solid black line and test samples in blue. The multimer image is depicted in the top right of each (A), von Willebrand factor antigen (VWF):Ag of 20 IU/dL densitometry LMWM 15.8%, IMWM 38%, HMWM 46.2%. (B) VWF:Ag of 15 IU/dL densitometry LMWM 19%, IMWM 35.8%, HMWM 45.2%. (C) VWF:Ag of 10 IU/dL densitometry LMWM 32%, IMWM 23.4%, HMWM 44.6%. (D) VWF:Ag of 5 IU/dL densitometry LMWM 30.8%, IMWM 23.4%, HMWM 45.8%