DOI: 10.14457/tu.the.2020.1004
title: Characterization of temperature-responsive poly (N-isopropylacrylamide-co-acrylamide) grafted culture surface

description: Poly (N-isopropylacrylamide) is a temperature-responsive polymer that has been widely utilized for tissue culture surfaces in cell sheet engineering. In this study, Poly (N-isopropyl acrylamide-co-acrylamide) (PNIAM-co-AM) is grafted onto tissue culture polystyrene substrate (TCPS) by using UV polymerization. The grafting density and thickness were controlled by UV irradiation time and spin coating speed to better achieve cell detachment. The morphology of the grafted surfaces was investigated by Atomic Force Microscopy (AFM). The results indicated that the surface become smoother as the spin speed increases. The water contact angle (WCA) measurement results show the wettability of grafted surface changed as the temperature changed without significantly change seen in ungrafted TCPS surface. The grafted amount of PNIAM-co-AM which varying UV irradiation time and spin speed was obtained between 1.68 to 2.03 μg/cm2 and thickness around 11-21 nm. Mouse preosteoblast MC3T3-E1 cells was reached 100 percent detachment with the samples prepared with 1.5 and 2-hours ultraviolet exposure times without the use of a poly (vinylidene difluoride) membrane. The detached sheet was in good condition, as indicated by Live/Dead stains.Along with this research, Spectroscopic Ellipsometry (SE) and Attenuated Total Reflection-Fourier Transform Infrared spectroscopy (ATR-FTIR) with synchrotron radiation were used to quantify the thickness and the grafting density, respectively. The amount of PNIAM-co-AM grafted surface was calculated from quantitative ATR-FTIR. The signal of peak area ratios at wavenumber 1,654 to 1,600 cm−1 (A1,654/A1,600) from known grafting densities were used to construct calibration curve expressed as linear equation (A1,654/A1,600 = 0.138σ). Spectroscopic ellipsometry was used to determine the thickness of polymer film. the mathematic dispersion models based on Cauchy equation and Lorentz equation were examined the best fit of the PS/PNIAM-co-AM layer by using uniqueness model testing. A combination of SE and ATR-FTIR was used to construct a prediction model of grafting density (σ) obtained from the film thickness (h). The linear model was expressed as h = 6.29σ + 2.24, with R2 = 0.93. To validate the predictive model, PNIAM-co-AM grafted TCPS surface with different thicknesses and grafting densities were examined. The result of predicted grafting density was nearly equal to the measured values. As a related study, the dehydration of copolymer in aqueous environment was studied by in-situ spectroscopic ellipsometry measurement. With a custom-made liquid cell, it is possible to study an ultrathin PNIAM-co-AM layer in a liquid environment, and to observe the polymer chain collapse and expand. The change of ѱ and Δ spectra, after reduced the temperature below 32°C, can be assumed as the change in thickness and refractive index of the copolymer which corresponding to the transition of the temperature-responsive PNIAM polymer. This technique can further development for constructing model of the thickness and optical properties of PNIAM-co-AM layer in a aqueous environment and has further implications for protein adsorption and desorption onto a temperature-responsive surface.