Surface engineering with poly(ethylene glycol) photolithography to create high-density cell arrays on glass

This manuscript presents a microfabrication-derived approach for controlling mammalian cell-surface interactions. Poly(ethylene glycol)-diacrylate (PEG-DA) was patterned, in a process analogous to photolithography, to manufacture high-density arrays of micrometer-scale PEG hydrogel wells on glass. Individual wells consisted of PEG walls and glass attachment pads; thus, as a result of the biological inertness of PEG, microwell patterning created a highly ordered biointerface with modulating-cell or protein-repellent properties. Fabricated hydrogel microstructures proved very effective in guiding and confining adhesion of transformed 3T3 fibroblasts and primary rat hepatocytes to defined regions on the glass substrate. PEG-patterned glass surfaces were selectively modified with collagen (type I) to induce hepatocyte attachment. Localization of the fluorescein-conjugated collagen within the glass attachment pads of PEG hydrogel microwells was visualized by fluorescence microscopy. Further surface analysis was performed by tapping mode atomic force microscopy conducted within individual PEG wells. Protein-modified regions inside the wells had a root-mean-square roughness of 1.13 ± 0.03 nm compared to 0.7 ± 0.04 nm for alkylsilane-treated regions lacking protein. The cell occupancy of 96.7 ± 1.9% within the arrays of 30 × 30 μm individual wells was demonstrated for 3T3 fibroblasts. At the same time, cells remained effectively isolated in the individual PEG microwells. Primary hepatocytes attached and became fully confluent within the collagen-coated PEG after 24 h of incubation. Each 30 × 30 μm well contained one to three hepatocytes. Cells patterned on the surface remained viable after 24 h of incubation.