In this work a novel and simple electrochemical sensor for the determination of (S)-warfarin based on an activated screen printed gold/Au nanoparticles/molecularly imprinted polymer electrode (ASPG/AuNPs/MIP) was introduced. Cyclic voltammetry, differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy were used to investigate the electrochemical behavior of (S)-warfarin at the modified electrode under optimum conditions. According to the results, ASPG/AuNPs/MIP showed a linear range for (S)-warfarin concentrations from 5.0 nM to 1.0 μM and a limit of detection of 1.0 nM by DPV. The proposed sensor was used for the determination of (S)-warfarin in commercial tablets, human serum and urine with satisfactory results.