Summary: Urea is one of the metabolic products of protein metabolism. The accurate determination of urea is important in glomerular filtration rate determination, renal function testing, and other biomedical applications. This research presents a simple fabrication of potentiometric urea biosensor based on a urease-immobilized polypyrrole (PPy). Thin film of PPy doped with hydroquinone monosulfonate (HQS) was electropolymerized on stainless steel supporting electrode. Urease was immobilized onto the PPy film by mean of physical adsorption. X-ray photoelectron spectroscopy (XPS) was performed to confirm the existence of urease on PPy surface. The PPy film immobilized with urease activity of 200 U/mL showed the sensitivity of 43.4 AE 3.3 mV/pUrea (r 2 > 0.99) in the urea concentration ranging from 0.5 to 10 mM (pUrea 2.0-3.3). The response time was less than 60 sec. The urease-immobilized PPy film offers a great potential in the use of conducting PPy film as an effective transducer to convert physiochemical change into an electronic signal, enhancing in performance of the biosensor.