Sb-doped ZnO microsphere film was fabricated by a carboxylate ion assisted hydrothermal route coupled with a post-calcination process. The structure, chemical composition and optical band gap of the Sb-doped ZnO microsphere film were characterized by X-ray diffraction, X-ray photoelectron spectroscopy, inductively couple plasma optical emission spectroscopy and UV-visible spectrophotometry, and compared with the un-doped ZnO microsphere film. The results suggest that the formation of zinc-antimony tartrate complex species during hydrothermal growth is the key to realize Sb-doped ZnO microstructures, and the present hydrothermal method with post-calcination is an effective way to dope Sb into ZnO. Furthermore, the Sb-doped ZnO microsphere film based electrochemical biosensor exhibits a good sensing performance for the detection of hydrogen peroxide, with a sensitivity of 271 mu A mM(-1) cm(-2) which is more than three times higher than that of the un-doped ZnO biosensor. (C) 2013 Elsevier B. V. All rights reserved.