Substituted polyaniline/chitosan (sPANI/Ch-H2SO4) composites were electrochemically synthesized in 0.10 M H2SO4 acid medium. The composites were characterized by electrochemical measurements, FTIR, SEM and AFM analyses. Anodic and cathodic current responses of substituted polyaniline/chitosan composites increased linearly with increasing the scan rate. SEM images of chitosan show smooth surfaces, whereas sPANI/Ch-H2SO4 composites show more porous surfaces which will be suitable for the immobilization of biomolecules. Glucose oxidase (GOD) was immobilized on to the surface of these composites by physical adsorption. The PNMANI/Ch-H2SO4/GOD electrode was utilized for glucose sensing for bigger concentrations range than other sPANI/Ch-H2SO4 composites and Ch/GOD by the amperometric method. The response current of PNMANI/Ch-H2SO4/GOD increases linearly with glucose concentration from 0.06 to 1.83 mM. The surface topography of the electrodes was studied using atomic force microscopy. PNMANI/Ch-H2SO4/ITO composite film electrode exhibited a relatively rough surface topography than Ch/ITO, which may facilitate the immobilization of biomolecules. (C) 2010 Elsevier Ltd. All rights reserved.