The attachment and spreading of fibroblast cells on a gold surface coated with fibronectin or ovalbumin were studied by a modified electric cell-substrate impedance sensor. In this system, cells were cultured in a well, equipped with a detecting gold electrode (surface area of 0.057 mm2) and a gold counter electrode (18 mm2). Based on a comprehensive theoretical framework, the impedance of the electrode-electrolyte interface and a cell layer was precisely obtained for frequencies ranging from 1 to 10 kHz. Surface concentrations of the protein adsorbed on the gold surface were determined by a surface plasmon resonance biosensor. The resistance change of the electrode-electrolyte interface at 4 kHz increased linearly with the number of fibroblast cells attached on the detecting electrode. The slope of the linear relationship appeared to depend on the type of coating protein. As the surface area occupied by the cells was also proportional to the cell number, the resistance change was in turn proportional to the area covered by the cells.