Cholesterol oxidase (ChOx) was physically entrapped in poly(3,4-ethylenedioxypyrrole) (PEDOP) to construct an amperometric cholesterol biosensor. The responses of the enzyme electrodes were measured via monitoring oxidation current of H2O2 at +0.7V in the absence of a mediator. Kinetic parameters, operational and storage stabilities, pH and temperature dependencies were determined. K-m, I-max and sensitivity (I-max/K-m) were calculated as 3.4 mM, 34 mu A cm(-2) and 10 mu A mM(-1) cm(-2), respectively. The minimum detectable substrate concentration was 0.4 mM and for a period of 20 days the biosensor showed the maximum relative activity. (C) 2008 Elsevier B.V. All rights reserved.