In this study, a new amperometric biosensor based on immobilized uricase was developed for the determination of uric acid. Polypyrrole-p-toluene sulphonate film was prepared on the surface of a platinum (Pt) electrode by electropolymerization of pyrrole in the presence of p-toluene sulphonate as an anionic dopant. Uricase enzyme were immobilized in polypyrrole-p-toluene sulphonate via the entrapment method. Determination of uric acid was performed by oxidation of enzymically generated H2O2 at 0.3 V. Some factors that affect response current were studied such as temperature, pH and substrate concentration. Operation stability and storage stability of the biosensor was determined. The biosensor retained 76.6% of its initial performance after 30 assays and it lost 58% of its initial performance after 60 days. Effects of interferants on the current response of the biosensor were examined. The performance of the biosensor was measured in serum of healthy individuals.