| AVS 59th Annual International Symposium and Exhibition | |
| Biomaterial Interfaces | Tuesday Sessions |
| Session BI+SS+AS-TuM |
| Session: | Biomolecules at Interfaces |
| Presenter: | D.G. Castner, University of Washington |
| Authors: | E. Tronic, University of Washington W. Thomas, University of Washington D.G. Castner, University of Washington |
| Correspondent: | Click to Email |
The clotting protein von Willebrand Factor (VWF) binds to platelet receptor glycoprotein 1ba (GP1ba) when VWF is activated, such as when VWF is exposed to a surface or is under high shear. However, the mechanism of surface activation is not known. This study characterizes function and adsorption behavior of the VWF A1 domain, which contains the GP1ba binding site. Surfaces tested are glass, polystyrene, and tissue culture polystyrene. Highest VWF A1- GP1ba binding is observed when A1 is adsorbed onto polystyrene, as measured by platelet rolling velocity in a parallel plate flow chamber assay. X-ray photoelectron spectroscopy (XPS) showed comparable A1 amounts are present on each surface, suggesting functional differences were not explained by differences in surface coverage. A1 surface structure was investigated using ELISA, time-of-flight secondary ion mass spectrometry (ToF-SIMS) and near-edge x-ray absorption fine structure (NEXAFS). Using monoclonal antibodies binding to a nonlinear epitope within A1, ELISA showed lower antibody binding for A1 adsorbed to polystyrene than to glass or tissue culture polystyrene. ToF-SIMS was used to identify differences in amino acid exposure, and NEXAFS showed different amide backbone ordering on the three surfaces. These studies demonstrate that the surface dependence of A1 function is likely due to differences in adsorbed surface orientation and/or conformation. This is an important consideration in in vitro models, where A1 is typically immobilized onto synthetic surfaces, and is also of interest for blood-contacting biomaterials. Additional studies have been done on A1 and two A1 mutants adsorbed on collagen coated tissue culture polystyrene. One mutant exhibits similar ELISA and ToF-SIMS results to the wild type A1, while the other mutant exhibits differences. This indicates that mutations in A1 can affect the conformation/orientation changes that result from A1 adsorption onto collagen.