A "redox-flexible" reagentless amperometric biosensor that can function either in oxidation or reduction and can cover large ranges of substrate concentrations has been developed for glucose measurements. Glucose oxidase (GOx) either uses O 2 as its natural electron acceptor and produces H 2 O 2 or uses artificial electron acceptors like ferricinium derivatives. To broaden the range of working potentials, peroxidase enzyme (HRP) was added to GOx, offering the alternative to correlate the glucose concentration to the reduction current of the ferricinium that results from H 2 O 2 oxidation of ferrocene. In such a configuration, the same ferrocene/ferricinium couple acts as a redox mediator for both enzymatic reactions involving GOx and HRP. Two ferrocenes were used in this study: Fc(CH 2 OH) 2 , substituted by electron-donor groups, and FcCOOH, substituted by an electron-withdrawing group. The rates of the reactions involved were determined, and the calibration curves in cathodic and anodic mode were drawn. The comparative study showed that for glucose measurement, the [FcCOOH]/[FcCOOH] + couple is best suited to act as mediator in the construction of a "redox-flexible" glucose biosensor.