BACKGROUND: Tetracycline (TC) is widely utilized in aquaculture as a broad-spectrum antibiotic with notable bactericidal properties. Nevertheless, the inappropriate or excessive application of TC may result in the presence of antibiotic residues in edible tissues, presenting significant risks to human health. In this study, a TC-sensitive electrochemical sensor was developed by modifying glassy carbon electrodes with gold nanoparticles (AuNPs)/graphene oxide (GO)/TC-templated molecularly imprinted polymer (MIP) composites. OBJECTIVE: A highly sensitive sensor for the detection of TC in milk. METHODS: The MIP, synthesized by thermal polymerization, acted as a selective recognition element and pre-concentrating agent for TC. To improve the conductivity of the MIP film and enhance the transfer of electrons across the electrode surface, AuNPs/GO composites were embedded in the MIP film. The morphology, structure, thermal stability, and electrochemical properties of the AuNPs/GO-MIP film were characterized through the utilization of scanning electron microscopy (SEM), FTIR spectroscopy, thermogravimetric analysis (TA), and electrochemical impedance spectroscopy (EIS). RESULTS: The modified electrode, featuring a composite film, exhibited a broad linear detection range (1-30 μg/L, 0.03-0.5 mg/L, and 0.5-20 mg/L), low detection limit (0.7 μg/L; S/N = 3), and high selectivity toward TC. CONCLUSIOS: The proposed sensor effectively quantified TC in milk. HIGHLIGHTS: In this study, AuNPs/GO composites were embedded to improve the conductivity of the composites; conductive MIP films were prepared and used as recognition elements and signal amplifiers. We present an electrochemical sensor for sensitive and selective detection of TC; the sensor can effectively quantify TC in milk.