In this study we have reported the design and development of a facile, sensitive, selective, and label-free electrochemical sensing platform for the detection of atrazine based on MWCNT-embedded ZnO nanofibers. Electrospun nanofibers were characterized using scanning electron microscope (SEM), transmission electron microscope (TEM), X-ray diffraction (XRD), X-ray photoelectron spectroscope (XPS), UV-Visible spectroscope (UV-VIS), and Fourier-transform infrared spectroscope (FTIR). Electrochemical properties of MWCNT-ZnO nanofiber-modified electrodes were assessed using electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). Binding event of atrazine to anti-atrazine antibody, which immobilized on nanofiber-modified electrode via EDC and NHS chemistry, was transduced with EIS. Due to high conductivity, surface area, and low bandgap of MWCNT-ZnO nanofibers, we have achieved the sensitivity and limit of detection (LoD) of sensor as 21.61 (KΩ μg−1 mL−1) cm−2 and 5.368 zM for a wide detection range of 10 zM–1 µM. The proposed immunosensing platform has good stability, selectivity, repeatability, and reproducibility, and are less prone to interference. © 2020, The Author(s).