Bimetallic Nanozyme-Assisted Immunoassay for the Detection of Acetamiprid in Vegetables

The ability of conventional enzyme-linked immunosorbent assays to satisfy the increasing demand for advanced immunoassays is limited by their reliance on natural enzymes, low sensitivity, and narrow detection range. The development of nanozyme-assisted immunoassays is a viable approach to overcoming these constraints. Here, we report a one-step bimetallic nanozyme-assisted indirect competitive immunoassay to detect the pesticide acetamiprid in vegetables. Litchi-like bimetallic gold and platinum (Au@Pt) nanozymes with high catalytic properties and excellent biocompatibility were synthesized for this assay. The Au@Pt nanozymes were simple and inexpensive to prepare, exhibited stability and adjustable catalytic activity, and were directly conjugated with antibodies as signal probes, without the use of horseradish peroxidase-conjugated Affinipure goat antimouse IgG (IgG-HRP). Acetamiprid was shown to compete with the antigen to bind the Au@Pt probes. Subsequently, Au@Pt probes, which exhibited peroxidase-like activity, were used to catalyze the oxidation of colorless 3,3′,5,5′-tetramethylbenzidine, demonstrating the easy detection of acetamiprid through colorimetric signal monitoring. The linear range, sensitivity, relative standard deviation range, and limit of detection of the immunoassay were 1.85–327.19 μg/L, 25.58 μg/L, 0.46–10.35%, and 0.78 μg/L, respectively, satisfying the detection requirements. In conclusion, the method exhibited high sensitivity, reproducibility, and a wide linear range for the detection of acetamiprid, while eliminating the need for IgG and simplifying the experimental procedure. This new indirect competitive immunoassay has great potential value for on-site sensitive detection of acetamiprid in vegetables.