Activated VWF present in VWD type 2B plasma. (A) Microtiter wells coated with AU/VWFa-11 (5 μg/mL) were blocked 30 minutes with 3% BSA, 0.1% Tween-20 in PBS. NPP (▪), plasma from healthy individuals (n = 9; □) and VWD type 2B plasma (n = 12; ▴, ▾) were diluted in PBS to obtain a concentration range (0.23-0.93 nM). After washing, wells were incubated 1 hour at 37°C with the diluted plasmas. Bound VWF was detected using HRP-conjugated polyclonal anti-VWF antibody. The concentration of VWF in the diluted samples was plotted against the measured OD 490 nm. The slope found for NPP was set to be 1. Arrows indicate the slopes found for 2 different VWD type 2B patients. (B) The activation factors were calculated and plotted in a scatter plot. Arrows indicate the activation factors calculated for patients 1 and 2 from panel A. The activation factor found for VWD type 2B patients was significantly higher than that for the healthy individuals (P < .001). Data represent the mean ± SD. (C) The activation factor calculated for 9 VWD type 2B patients was plotted against the thrombocyte counts in these samples and a correlation was found to be significant (P < .003; R2 = 0.7401).
