Nanoporous Al2O3 membranes are employed in the development of amperometric biosensors for the immobilization of the biologically active elements. Anodized alumina membranes with the desired dimensions were prepared by controlled anodization of high purity aluminium in an acidic solution that slowly dissolves the formed aluminium oxide. The anodizing voltage controls the pore size and pore density, whereas the thickness is determined by the amount of transferred charge. This technique originates in the membrane an asymmetric structure of cylindrical pores of uniform diameter arranged in a hexagonal array perpendicular to the membrane surface. Such structure allows the uptake of relatively higher amounts of glucose oxidase (GOx) enzyme in a thin film. The preparation of the active phase of the biosensors is completed by procuring an external coating to the membrane surface with a natural polysaccharide, chitosan, which avoids the GOx leaching and provides a cage of high surface area inside the pores where the entrapped enzyme remains active. The aim of this study is to evaluate the influence of membrane dimensions on the biosensor response. (c) 2005 Elsevier B.V. All rights reserved.