The aim of this research was to develop a simple, selective and sensitive aptasensor for detection of Pb2+ based on hairpin structure of complementary strand (CS) of aptamer (Apt). Screen printed carbon electrode (SPCE) was electrodeposited by gold nanoparticles to be used as the substrate for the immobilization of Apt-CS. Moreover, gold nanoparticles (AuNPs) and thionine were added to increase the sensitivity of the aptasensor. The morphology of SPCE and AuNPs modified SPCE was investigated using scanning electron microscopy (SEM). The SEM results showed that the AuNPs were homogeneously distributed on the surface of SPCE. The electrochemical performance of aptasensor was evaluated by means of cyclic voltammetry (CV) and differential pulse voltammetry (DPV) measurements. The electrochemical results indicated that peak current due to the hairpin structure formation of CS decreased in the presence of Pb2+. However, in the absence of Pb2+, peak current was enhanced because of the combination of thionine/AuNPs-CS. The designed aptasensor showed a high selectivity toward Pb2+, wide linear range (1-40 nM), low detection limit (374 pM), acceptable reproducibility and good long-term stability.