Polyaniline (PANi) microtube arrays were synthesized by a secondary-template (fir wood and ZnO microtube array) method and confirmed by scanning electron microscopy (SEM). High magnification SEM observation revealed that the PANi microtube arrays were formed by nanospheres and nanorods. X-Ray diffraction, Fourier transform infrared spectroscopy and cyclic voltammetry were used to characterize the resultant PANi microtube arrays which were subsequently used as electrodes for glucose oxidase (GO) immobilization and the detection of glucose. Compared with the traditional PANi powder electrode, the bioelectrocatalytic activation of GO immobilized PANi microtube array electrode toward the oxidation of glucose has been significantly enhanced. The excellent properties, low cost, facile preparation and good biocompatibility of the PANi microtube arrays set a bridge between proteins/enzymes and electrodes, providing analytical access to a large group of enzymes for a variety of biosensor applications.