Analysis of the capacity of the different human-murine VWF chimeras to promote platelet adhesion in flow. Blood was collected from mice 1 day after hydrodynamic injection with pCDNA6 encoding muVwf, huVWF, or one of the chimeric variants. PPACK-anticoagulated whole blood was incubated with rhodamine 6G to fluorescently label platelets and perfused over collagen-coated glass coverslips (flow rate 2500 seconds−1) for a period of 1 minute. Unbound platelets were removed by subsequent perfusion with HEPES-Tyrode buffer. Thrombus formation was then visualized via image acquisition using Metamorph Version 7.0rl software. Shown are representative images for muVwf, huVWF, huVWF H1326R, and huVWFmuA1. Thrombus formation was quantified using ImageJ Version 1.44 software to calculate percentages of platelet surface coverage. Data represent the mean ± SEM of 3 independent perfusions, with 7-19 images being analyzed for each perfusion. Statistics were performed using 1-way ANOVA followed by the Dunnett multiple comparison test. The various constructs were compared with muVwf, and all except huVWFmu (1326-1333; 1370-1385) and huVWFmuA1 led to *P values < .05.
