Abstract A simple, selective, highly sensitive, and label-free electrochemical immunosensor for atrazine detection was developed by immobilizing gold nanoparticles (GNPs) on the gold electrode surface. The modified gold electrode exhibited good electrochemical activity. The high sensitive detection was achieved by increased the surface area of work electrode resulting in capturing more anti-atrazine monoclonal antibodies. The whole immunosensor fabrication process was characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) with the use of ferricyanide as an electrochemical redox indicator, respectively. The signal was triggered based on the interaction between anti-atrazine monoclonal antibody and atrazine via differential pulse voltammetry (DPV). Under the optimal conditions, a limit of detection for atrazine as low as 0.016 ng/mL was obtained in buffer, and the linear working range was between 0.05 ng/mL and 0.5 ng/mL. The recovery of 95.5–119.8% was obtained in maize. The proposed immunosensor has acceptable stability and reproducibility, thus could be applied to other pesticides analysis of crop samples in practice.