Discrimination of the S-isomer of ofloxacin using nanozyme-enhanced immunochromatographic analysis based on a secondary antibody-modified Au@Pd nanozyme

Ofloxacin, a fluoroquinolone antibiotic, is a significant contaminant in food products. Its levorotatory form (S-OFL) exhibits a bactericidal activity considerably higher than the dextrorotatory form (R-OFL) and is commonly employed in veterinary medicines. In this work, a highly sensitive immunochromatographic analysis (ICA) method for biologically active S-OFL was developed. A bimetallic core@shell Au@Pd nanozyme was synthesized to serve as the label. Thanks to its peroxidase-like catalytic activity, the nanozyme enhances the color signal on test strips, thereby improving assay sensitivity. The instrumental limit of detection and cutoff values were determined to be 1.4 pg/mL and 0.1 ng/mL, corresponding to 60- and 10-fold improvements compared with conventional gold nanoparticle-based ICA. The cross-reactivity with R-OFL was only 6.3%, demonstrating the high specificity toward target isomer. The ICA was successfully applied to detecting S-OFL in milk, achieving recoveries of 77–99%. The entire nanozyme-enhanced analysis can be completed within 20 min. This method shows potential for monitoring various toxicants in food products.