Honeycomb-patterned polymer films are formed upon self-organization of water microspheres stabilized by amphiphilic polymers. The honeycomb films can work as cell culture substrates. The films composed of heparin-cationic lipid complex, however, gradually lost the honeycomb structure when immersed in culture medium. Crosslinking of the films was carried to stabilize the honeycomb pattern. A bisazido derivative was used as a crosslinker connecting the constituent polymer of the honeycomb films. Morphological study of the immersed films revealed that the crosslinked films maintained the honeycomb structure in the phosphate buffer solution.