<p>The rapid and sensitive detection of trace antibiotics in environmental and biological samples is crucial for food safety and public health. This study presents a novel electrochemical aptasensor based on a porous covalent organic frameworks/graphene oxide (namely COF/GO) hybrid material for selectively detecting penicillin (PCL). A specific aptamer was immobilized on the modified electrode surface for high-affinity recognition of PCL. The binding event between the aptamer and PCL induces a measurable change in the electrochemical signal, which is quantitatively analyzed using differential pulse voltammetry (DPV). The developed aptasensor demonstrated excellent performance with a wide linear range from 0.01 ng mL^− 1 to 10.0 ng mL^− 1 and an ultralow detection limit of 2.9 pg mL^− 1. The sensor exhibited high selectivity against interfering substances and good stability for storing 8 days. This work highlights the potential of porous COF/GO composite-based electrochemical aptasensors as a powerful tool for real-time monitoring of antibiotic residues in complex matrices.</p> Graphical Abstract <p></p>