For preparing a thermally modulated biointerface that separates cells without the modification of cell surfaces for regenerative medicine and tissue engineering, poly(N-isopropylacrylamide-co-butyl methacrylate) (P(IPAAm-co-BMA), thermoresponsive hydrophobic copolymer brushes with various BMA composition were formed on glass substrate through a surface-initiated atom transfer radical polymerization (ATRP). Characterization of the prepared surface was performed by X-ray photoelectron spectroscopy (XPS), attenuated total reflection Fourier transform infrared spectroscopy (ATR/FT-IR), and gel-permeation chromatography (GPC) measurement. Prepared copolymer brush surfaces were characterized by observing the adhesion (37 degrees C) and detachment (20 or 10 degrees C) of four types of human cells: human umbilical vein endothelial cells (HUVECs), normal human dermal fibroblasts (NHDFs), human aortic smooth muscle cells (SMCs), and human skeletal muscle myoblast cells (HSMMs). HUVECs and NHDFs exhibited their effective detachment temperature at 20 and 10 degrees C, respectively. Using cells' intrinsic temperature sensitivity for detachment from the copolymer brush, a mixture of green fluorescent protein (GFP)-expressing HUVECs (GFP-HUVECs) and NHDFs was separated.