The widespread use of toltrazuril (TOL) as a low-toxicity anticoccidial agent in livestock production necessitates sensitive monitoring of its persistent metabolite, toltrazuril sulfone (TOS), which exhibits hepatotoxic and reproductive risks at subchronic exposure levels. Here, we proposed a novel electrochemical immunosensor for the ultrasensitive detection of TOS in eggs, utilizing a decorated amino-functionalized cerium metal-organic framework-gold nanoparticle (NH₂-Ce-MOF/AuNPs) nanocomposite. The NH₂-Ce-MOF was synthesized via a hydrothermal crystallization of (NH₄)₂Ce(NO₃)₆ and 2-aminoterephthalic acid, exhibiting exceptional stability and an expansive surface area. Compared to conventional electrodes, NH₂-Ce-MOF-modified electrodes promoted AuNPs nucleation and provided more active sites for HAuCl₄ deposition, while also enhancing antibody binding, thereby improving the detection sensitivity of electrochemical immunosensors. Subsequently, electrodeposited HAuCl₄ enhanced both electron transfer kinetics (charge transfer resistance decreased by 68% to a bare electrode) and bioreceptor loading capacity. Electrochemical evaluation demonstrated superior sensor performance, achieving a wide linear range of 0.05 ng/mL–100 ng/mL (R²=0.998) with an ultralow limit of detection of 33.2 pg/mL (S/N=3), representing a 3-fold improvement over conventional enzyme-linked immunosorbent assays. The electrochemical immunosensor exhibited excellent selectivity and reproducibility (relative standard deviation<10%). This platform establishes a robust foundation for TOS residues and other hazardous contaminant monitoring in complex food matrices.

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