A novel electrochemical sensor for paraquat (PQ) detection based on molecularly imprinted polymer (MIP) membranes on a glassy carbon electrode (GCE) modified with Au nanocrosses-chitosan (AuNCs-CS) was constructed. P-Aminothiophenol (p-ATP) and 4,4′-bipyridine template were assembled on the surface of the modified GCE by the formation of Au-S bonds and hydrogen-bonding interactions, followed by polymer membrane formation by the electropolymerization in a polymer solution containing p-ATP, HAuCl 4 , tetrabutylammonium perchlorate (TBAP), and the template molecule 4,4′-bipyridine. The asconstructed molecularly imprinted sensor (MIP-AuNC-CS) was characterized by differential pulse voltammetry (DPV), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). This is the first time that molecularly imprinted polymer technology has been integrated with a modified AuNCs-CS to electrochemically detect PQ. The linear response of the MIP-AuNC-CS sensor was in the range from 1 × 10 −14 to 1 × 10 −10 mol L −1 , and the limit of detection was 2.3 × 10 −15 mol L −1 . This sensor showed high-speed real-time detection capability, low sample consumption, high sensitivity, low interference, and good stability characteristics, and was proven to detect PQ.