:Dodecylbenzene sulfonate (DBS)-doped polypyrrole (PPY) conducting polymer films were electrochemically deposited onto the indium-tin-oxide (ITO)-coated glass plates in aqueous medium. The enzyme cholesterol oxidase (ChOx) was immobilized on these DBS-PPY films by a physical adsorption technique. These ChOx-immobilized DBS-PPY films were characterized by ultraviolet-visible and Fourier transform infrared spectroscopy. The enzyme activity studies indicate that similar to 40% of ChOx leaches out from the ChOx/DBS-PPY film. The ChOx activity in the ChOx/DBS-PPY film was assayed as a function of cholesterol concentration. The results of amperometric measurements conducted on ChOx/DBS-PPY/ITO film show linearity over the range 2-8 mM of cholesterol solution. The ChOx/DBS-PPY/ITO electrodes exhibit a response time of 30 s and are stable for similar to3 months at 4 degreesC.