A composite film that contained CeO2-ZrO2 hollow nanospheres and chitosan was deposited on a gold electrode using a one-step electrodeposition based on the pH-dependent solubility of chitosan. Electrochemical characterization experiments showed that the electroactive surface of the modified electrode was significantly enhanced by the introduction of the CeO2-ZrO2 nanospheres into the chitosan gel during electrodeposition. Furthermore, an electrochemical DNA biosensor was fabricated based on the synergistic affinity of CeO2-ZrO2 and chitosan in the nanocomposite film to probe DNA. DNA hybridization assays suggested that the target DNA could be quantified over a wide dynamic range of 1.63 x 10(-13) M-1.63 x 10(-8) M with a detection limit of 1.0 x 10(-13) M using methylene blue as an electrochemical indicator. This work describes a simple strategy for the preparation of a stable and conductive interface for electrochemical detection of DNA hybridization and opens a path for the application of CeO2-ZrO2 nanospheres and chitosan in DNA electrochemical biosensing analysis. (C) 2011 Elsevier Ltd. All rights reserved.