Here we report a novel self-aligned process for gold nanoparticle (AuNP) decorated polypyrrole (Ppy) biosensing microelectrodes, where the AuNP selectively electro-grows out of the Ppy. The self-aligned process allows "on chip" AuNP preparation and deposition in one step, saving time and cost. The Au nanoparticles were galvanostatically deposited, yielding a random array of nanoparticles under specific conditions. Astudy of the AuNP size and coverage as function of deposition time and current density was accomplished using environmental scanning electron microscope (ESEM). AuNP array with less than one nano-particle monolayer thickness could be reproducibly maintained keeping the device mechanical flexible. This yields electrochemical behavior that was similar to that of bare gold electrode as was determined by cyclic voltammetry in biological solution with the Fe3+/Fe2+ redox couple. The electrochemical impedance spectroscopy (EIS) study of such AuNP electrodes was performed and modeled taking into consideration the composite nature of the electrode. The fabricated electrode with 10mA/cm(2) AuNP deposited Ppy electrode demonstrated good sensing ability of alkaline phosphatase enzyme by cyclic voltammetry and chronoamperometry at 0.22 V (vs. Ag/AgCl quasi reference electrode in 0.1M PBS pH 7.2). The average current response of the AuNP/Ppy was found to be 30% higher than the standard Au electrode. (C) 2017 The Electrochemical Society. All rights reserved.