Preparation of self-organized micro-patterned polymer films having cell adhesive Ligands

This article describes novel three methods for micro-patterning of cell adhesive ligands by using the self-organized honeycomb-patterned structure formed by the simple cast method. A first method is a direct preparation of a patterned film by casting an amphiphilic polymer containing lactose residue which is one of cell adhesive ligands. A benzene solution of the amphiphilic polymer was cast at high humidity on a glass substrate. Atomic force microscopy (AFM) observation of the film showed that a honeycomb pattern with microporousness with as large as micrometer size in diameter was formed. The film was immersed into an aqueous fluorescence-labeled lectin solution to investigate the distribution of lactoses on the patterned film. Consistence of a fluorescence image of the lectin bound film with the honeycomb pattern showed that the lactose residues were existed not at the holes but at the rims of the honeycomb-patterned film. A second method is to immobilize gelatin, which is one also one of cell adhesive ligands, on the honeycomb-patterned film by chemical reaction. A honeycomb-patterned film was prepared from chloroform solution of an amphiphilic polymer containing reactive succinimide ester groups, and then the film was immersed into an aqueous fluorescence-labeled gelatin solution to introduce gelatin on the film surface. Immobilization of gelatin onto honeycomb-patterned film was confirmed by the fluorescence microscope. A third method is another way to introduce gelatin onto the honeycomb film by the specific avidin-biotin interaction. A honeycomb-patterned film was prepared from amphiphilic polymer containing biotin residues and dodecyl groups, and then the film was immersed into a avidin solution and a biotinylated fluorescence labeled gelatin solution successively. By the fluorescence microscopic observation of the film, gelatin was confirmed to be immobilized at the rims of the honeycomb pattern via the avidin-biotin interaction. Cell culture was performed on the gelatin immobilized patterned film prepared by second method. Bioactivity of gelatin immobilized honeycomb-patterned film was confirmed by adhesion of cell onto the film.