Electron Microscopy Visualization of Vitronectin Adsorbed on COOH and NH₂ Functionalized Surfaces: Distinctive Spatial Alignment and Regulated Cellular Responses

Abstract Adsorption of proteins associating with their conformational changes plays crucial roles in regulating biomaterial–cell interactions and consequent tissue responses to implanted biomaterials. This study reports direct visualization of typical serum protein, vitronectin, one of the key adhesive proteins that participate in mediating cell behaviors, upon adsorption on typically designed surfaces. Carbon films with their surfaces being plasma grafted functional groups COOH and NH₂ are used as the model substrata for this study. Negative‐staining electron microscopy technique is employed for visualizing the adsorbed protein and 2D image classification is made and interpreted. Results show that adsorbed vitronectin tends to form multimer aggregate on the COOH‐grafted surfaces, exposing extensively its cell‐binding RGD (arginine‐glycine‐aspartic acid) motif for enhanced cell adhesion. The adsorbed vitronectin on the NH₂‐grafted surface forms dimer aggregate with the binding sites being enwrapped. The COOH‐grafting triggers enhanced expressions of ITGA5, ITGAV, ITGB1, and ITGB3 of the adhered cells and this is likely attributed to the special spatial alignment of vitronectin upon adsorption. The conformational information of adsorbed vitronectin gained from the single particle electron microscopy analyses would shed light on design and construction of appropriate biomaterials surfaces for desired cellular behaviors.