Zearalenone (ZEN) is an important non-steroidal estrogenic mycotoxin produced by fusarium fungi species. A number of methods based on enzyme-linked immune assays (ELISA) and chromatographic approaches are available for the detection of ZEN. In this research, ZEN conjugated by bovine serum albumin (BSA) was immobilized on the surface of the screen-printed carbon electrode (SPCE) followed by the subsequent addition of anti-zearalenone monoclonal antibody (ZEN-MAb), alkaline phosphatase (ALP)-labeled secondary antibody, and 1-napthyl phosphate (1-NP) as a substrate. This immunosensor had good specificity, selectivity, reproducibility, and sensitivity for ZEN (LOD of 0.25 ng mL−1) when employed for quantification based on differential pulse voltammetry (DPV). This immunosensor yielded good recovery values in the range of 89–97% against real matrices (i.e., beer and wine samples) after spiking. This novel immunosensing principle can be extensively applied for the development of advanced sensing techniques for various analytes. © 2018