Temperature responsive poly(N-isopropylacrylamide-co-styrene) nanofilms for non-enzymatic cell sheet harvesting

Abstract

In this study, temperature-responsive cell culture substrates with non-enzymatic cell sheet detachment attributes in response to temperature reduction were prepared. Poly(N-isopropylacrylamide) (PNIPAAm) with a physiologically proximal lower critical solution temperature (LCST) induces a temperature responsivity to the substrate, and polystyrene was used to regulate the hydrophobicity of the substrate. Different compositions of poly(NIPAAm-co-styrene) were synthesized by free radical polymerization. The resulting copolymers were characterized using Fourier transform infrared spectroscopy (FTIR) and proton nuclear magnetic resonance (1HNMR) spectroscopy. The copolymer composition was calculated from 1HNMR results. The molecular weight and molecular weight distribution were measured by gel permeation chromatography (GPC). The nanofilms of the prepared copolymers were physically coated by spin-coating technique on tetramethyl orthosilicate (TMOS) pre-coated glass slides. Attenuated total reflection-FTIR (ATR-FTIR) and field emission scanning electron microscopy (FESEM) analyses showed that the spin coating procedure was performed successfully, and the change in the water contact angle (WCA) in response to temperature alteration indicated the temperature-responsive behavior of the polymer layers. L929 mouse fibroblast cell line was cultured on nanofilm-coated glass slides until a confluent cell sheet covered the entire surface. An almost integrated cell sheet was harvested by temperature reduction from 37 °C to 20 °C. The results revealed that copolymer composition is vital in harvesting contiguous cell sheets. This fabrication method, compared to others such as plasma polymerization and surface-initiated atom transfer radical polymerization (SI-ATRP), is very cost-effective and applicable for large-scale cell culture investigations.