Self-assembled Monolayer-Based Surface Modification of Stainless Steel for Oriented Antibody Immobilization: Towards Improved Stent Biocompatibility
Keywords: antibodies
endothelial progenitor cells
stainless steel
stent
trichlorosilane monolayer

Stenting is a common surgical procedure for the widening of arteries narrowed by atherosclerosis. The stent is an expandable cylindrical scaffold that is implanted within the lumen of arteries and is commonly constructed from stainless steel. Despite the benefits of stenting, post-stent re-narrowing of arteries due to neointimal hyperplasia and biocompatibility associated challenges continues to limit the efficacy of bare metal stents. In order to mitigate these problems, drug-eluting stents have been developed to pharmaceutically interrupt these processes. While successful, concerns regarding late stent thrombosis and delayed healing continue to surround this drug-based approach. The primary objective of this thesis is the development of a proof-of-concept stainless steel coating that could eventually be applied to stent technology for the reduction of neointima formation and stent surface fouling for improved healing and decreased use of therapeutic agents. The overall approach attempts to covalently attach oriented antibodies to 316L stainless steel surfaces through a crosslinking self-assembled monolayer (SAM). The employed linker for SAM formation, termed TUBTS, is a novel silane recently developed by our research group. It is a bifunctional molecule that bears a trichlorosilyl anchoring group and a thiol-reactive head function. The antibody of interest is one capable of binding endothelial progenitor cells (EPCs). Attachment of these cells to the stent surface confers the antifouling properties to the coating as these cells have been found to participate in the re-endothelialization of damaged arterial vessels and the inhibition of neointima formation. Successful cell attachment requires the antibodies to be properly oriented on the SAM surface. To meet this requirement, the antibodies were modified with a thiol moiety opposite to the paratope such that the thiol reactivity of TUBTS could be used to orient the antibody as it is immobilized to the SAM. TUBTS SAMs were successfully formed on steel and used to immobilize intact antibodies and their Fabʹ fragments. While covalent attachment could not be directly proven, the immobilized proteins were found to be strongly bound and both the intact antibodies and Fabʹ fragments were shown to retain their antigen binding capabilities upon immobilization. Another key finding was the observation that the Fabʹ fragment strategy would constitute a preferential option to that involving intact antibodies when considered in the context of in vivo capture of EPCs in stent applications. From the foundational work presented here, EPC binding to both forms of biofunctionalized TUBTS-modified steel surfaces is now being investigated. In anticipation of potential problems in this on-going work, a strategy for increasing the spacing between the TUBTS-modified surface and immobilized Fabʹ fragments is also presented alongside suggested methods for assessing the mode of Fabʹ fragment attachment.