A nanostructured electrochemical sensor was developed to establish an innovative way to measure ropinirole and levodopa in real pharmaceutical and biological samples, which offers a high selectivity derived from the stable carbon nanotubes-Nafion polymer film. The surface morphology of the modified electrodes was characterized by scanning electron microscopy. The effect of surface modifications, supporting electrolyte, amount of carbon nanotubes suspension, accumulation time and potential were investigated. A sensitive electroanalytical methodology for the simultaneous determination of both drugs co-administrated in advanced Parkinson's disease patients using adsorptive stripping square-wave voltammetry is presented. Under the optimized conditions, ropinirole and levodopa gave a linear response in the range 1 x 10(-7)-1 x 10(-5) M and 2.5 x 10(-7)-1 x 10(-5) M with detection limits 1.6 x 10(-8) M and 5.2 x 10(-8) M, respectively. The method was successfully utilised for their quantification in human serum samples and good recoveries were obtained without interference from endogenous dopamine, uric and ascorbic acid. In addition, the proposed sensor was successfully applied in the independent determination of ropinirole and levodopa content in pharmaceutical formulations, whose accuracy was attested by good agreement of the results with those, obtained using high performance liquid chromatography. The proposed sensor is characterized by high sensitivity and reproducibility, simple fabrication procedure, easy handling, resistance against surface fouling and low cost. (C) 2014 Elsevier B.V. All rights reserved.