Abstract Substratum surface morphology plays a vital roles in cellular behavior. Here, we characterized adsorption of fibronectin (Fn) as a typical cell adhesion protein onto honeycomb-patterned films made of poly(ε-caprolactone) (PCL) by using atomic force microscopy (AFM) and confocal laser scanning microscopy (CLSM). In order to determine how cells adhere to a honeycomb-patterned film, focal adhesion of cardiac myocytes (CMYs) and endothelial cells (ECs) on the films were studied by using fluorescence labeling of vinculin. Fn adsorbs around the pore edges to form ring-shaped structures. CMYs and ECs adhere onto the honeycomb-patterned films at focal contact points localized around pore edges distributed over the entire cellular surface. The focal contact points on the honeycomb-patterned films correspond well with the adsorption sites of Fn. We suggest that the cell response to honeycomb-patterned films is associated with the adsorption pattern of Fn on the film.