Thermoresponsive anionic copolymer brush-grafted surfaces for cell separation

Cell separation methods that do not require surface modification of cells are needed in tissue engineering and regenerative medicine. We developed thermoresponsive anionic polymer brushes for cell separation without modification of the cell surfaces. Copolymer brush poly(N-isopropylacrylamide-co-N-tert-butylacrylamide-co-tert-butyl acrylate, P(NIPAAm-co-tBAAm-co-tBA), was prepared on a cover glass plate through activator regenerated by electron transfer atom transfer radical polymerization (ARGET-ATRP). The tert-butyl group of the copolymer brush was then deprotected and a P(NIPAAm-co-tBAAm-co-acrylic acid (AAc)) brush-modified glass surface was obtained. ARGET-ATRP synthesis achieved polymers with low polydispersity. The negative surface charge of the polymer brush-modified substrates was evaluated using zeta potential measurements and the phase transition temperature of the polymer was modulated between 37-20 °C to perform cell adhesion and detachment, respectively. The adhesion and detachment behavior of cells used in cardiovascular tissue engineering on the thermoresponsive anionic polymer brushes was investigated. Normal human umbilical vein endothelial cells (HUVEC) exhibited prompt detachment from the thermoresponsive anionic polymer brush surfaces. In addition, normal human aortic smooth muscle cells (SMC) exhibited relatively high adhesion on thermoresponsive anionic polymer brush-modified surfaces compared with those modified with thermoresponsive polymer brushes without anionic groups. By utilizing the difference in the cell adhesion and detachment properties of the cell types, a mixture of HUVEC and SMC was separated simply by altering the applied temperature. This result indicated that the prepared thermoresponsive anionic polymer brush-modified glass surface could be used as a tool for the separation of cells in cardiovascular tissue engineering.