The immobilized cell using self-assembled synthetic oligopeptide was applied to the biological toxicity detection of environmental pollutant. Thin films based on cysteine-terminated synthetic oligopeptides were fabricated for the immobilization of Escherichia coli O157:H7 on gold (Au) substrate. Layer formation and immobilization of E. coli O157:H7 were investigated with surface plasmon resonance (SPR) and atomic force microscopy (AFM). Experimental results showed that the thin film of cysteine-terminated synthetic oligopeptide was successfully fabricated and it could be applied for the immobilization of E. coli O157:H7. The attached living cell was exposed to toxic chemical such as phenol, which induced the change of SPR angle. As the exposed concentration of phenol was increased, the change of plasmon resonance angle was increased, which indicates the decrease of cell viability. The detection limit based on SPR was determined as 5 ppm. The proposed cell immobilization method using self-assembly technique can be applied to construct the cell microarray for the diagnosis, drug detection, and on-site monitoring.