The aim of this work was to develop engineered coatings for protein and cell patterning on a surface. Cell patterning is important for biomedical applications such as single-cell studies, cell microcolonies arrays for high throughput drug screening, and the growth of geometrically controlled cell co-cultures for tissue engineering.udIn this work, the patterning of cells relied on the controlled positioning of protein domains on a surface, on which the cells could adhere. This was done by dewetting a protein-repellent polymer film from a protein-adsorptive one. Dewetting is the process by which an unstable thin liquid film (such as a polymer over its glass transition temperature) spontaneously breaks up, resulting in the formation of holes. The duration of dewetting controls the dimensions of the holes, from tens of nanometers to tens of microns. By tuning the thickness of the films and the molecular weight of the polymers it is possible to vary the type of pattern that is obtained. The result of the dewetting was a chemically and topographically patterned coating. Proteins in contact with such surface could only adsorb inside the dewetted holes, where the adsorptive polymer was exposed.udThe first system investigated was dewetted poly(N-vinylpyrrolidone) (PNVP) on polystyrene (PS). PNVP was found to cross-link upon annealing, as well as dewetting from PS. Insoluble cross-linked PNVP films were characterized by neutron reflectometry, infrared spectroscopy and ellipsometry, and found to be stable in water for many days, resistant to harsh solvents, and excellent in repelling proteins. The hole growth observed during concurrent dewetting and cross-linking was fully characterized by time elapsed optical microscopy, and a model was developed to predict it. The pattern obtained by dewetting PNVP could be controlled by selecting the appropriate annealing temperature, in order to tune the ratio between the rates of dewetting and cross-li nking.udThe PNVP/PS architecture was improved by substit! uting th e PNVP film with a functional polymer brush, in order to achieve a more versatile system. A polymer was designed so as to be able to dewet from PS, as well as bear initiators for the grafting of a polymer brush. This macroinitiator was synthesised by reversible addition-fragmentation chain transfer polymerization (RAFT). A protein-repellent poly(poly(ethylene glycol)methyl ether methacrylate) (poly(PEGMA)) brush was grafted from the dewettable macroinitiator film using activators generated by electron transfer atom transfer radical polymerization (AGET ATRP). This type of polymerization allows growing brushes of controlled thickness, with “living” ends that may easily be post-functionalised with simple chemical reactions, to interact selectively with different biological molecules or cells. The grafting process was investigated by ellipsometry, size exclusion chromatography and X-ray photoelectron spectroscopy. The functional patterned coatings developed were able to effe ctively immobilise extracellular matrix proteins and cells in selected areas of the surface, as shown by fluorescence microscopy and atomic force microscopy. The cells could spread on the surface, showing good viability.udThe patterned coatings here described could be prepared on non-flat and large objects, offering a simple and cheap alternative to other patterning techniques, such as photolithography and micro-contact printing, and opening exciting prospects in biomedical applications.
展开▼
