Investigation of the Morphology of Recombinant Von Willebrand Factor by Two High-Resolution Microscopic Techniques

Abstract 1409 Von Willebrand factor (VWF) is the largest known glycoprotein circulating in human blood plasma and is composed of a series of multimers with molecular weights ranging from 600 to 20,000 kDa or even more. In this study, we investigated the morphology of recombinant VWF (rVWF) and compared it to that of plasma-derived VWF (pdVWF) by tapping mode atomic force microscopy and transmission electron microscopy. Tapping mode atomic force microscopy showed that both rVWF and pdVWF contain globular and stretched domains. Mean chain lengths of the filaments and diameters of the core globular domains were determined and analyzed on a statistical basis. About 70% of the rVWF and pdVWF molecules were 100–300 nm long. The portion of very long molecules (>300 nm) was only slightly greater in rVWF than in pdVWF (20% versus 18%). The diameters of the globular core structures were in the range of 12 to 30 nm for both types of VWF. Inspection of a purified rVWF dimer revealed a similar range for the globular domain (14-32 nm). Upon exposure to high shear stress, a dramatic conformational change was observed for rVWF, just as has been reported for pdVWF. Investigation of negatively stained preparations of rVWF by transmission electron microscopy showed a highly organized structure. The heterogeneity in size observed was as expected for a polymeric VWF that consists of a mixture of multimers of varying sizes. The micrographs also revealed the presence of coiled and elongated structures. Analysis of pdVWF led to similar results, which were also in agreement with data from the literature. The results confirmed that the morphology of rVWF is similar to that observed for VWF purified from human normal plasma. Disclosures: Rottensteiner: Baxter Innovations GmbH: Employment. Seyfried: Baxter Innovations GmbH: Employment. Friedbacher: Vienna University of Technology: Consultancy. Allmaier: Vienna University of Technology: Consultancy. Ilk: University of Natural Resources and Applied Life Sciences: Consultancy. Sleytr: University of Natural Resources and Applied Life Sciences: Consultancy. Ehrlich: Baxter Innovations GmbH: Employment. Turecek: Baxter BioScience: Employment.