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Abstract: A novel film of chitosan-gold nanoparticles is fabricated by a direct and facile electrochemical deposition method and its application in glucose biosensor is investigated. HAuCl4 solution is mixed with chitosan and electrochemically reduced to gold nanoparticles, which can be stabilized by chitosan and electrodeposited onto glassy carbon electrode surfaces along with the electrodeposition of chitosan. Then a model enzyme, glucose oxidase (GOD) is immobilized onto the resulting film to construct a glucose biosensor through self-assembly. The resulting modified electrode surfaces are characterized with both AFM and cyclic voltammetry. Effects of chitosan and HAuCl4 concentration in the mixture together with the deposition time and the applied voltage on the amperometric response of the biosensor are also investigated. The linear range of the glucose biosensor is from 5.0×10−5 ∼1.30×10−3 M with a Michaelis–Menten constant of 3.5 mM and a detection limit of about 13 μM. [Copyright &y& Elsevier]