Development of a colloidal gold immunochromatographic strip for the rapid detection of pefloxacin in grass carp with a novel pretreatment method

A rapid colloidal gold immunochromatography assay (GICA) for the detection of pefloxacin (PEF) was established and optimized. The anti-PEF monoclonal antibody (mAb) was used to target PEF as a colloidal gold-mAb conjugate. The mAb belonged to the IgG2b subtype, lambda light chain, the affinity constant (Ka) was 5.21 × 10⁹ L·mol^-1, and its half maximal inhibitory concentration (IC₅₀) was 0.23 ng·mL^-1. No obvious cross-reactivity (CR) was observed with other common fluoroquinolone antibiotics, including ciprofloxacin (CIP), norfloxacin (NOR), lomefloxacin (LOM) and ofloxacin (OFL). The visual limit of detection (vLOD) of the optimized GICA was 2 ng·g^-1 under the conventional pretreatment method, and the assay was completed in 15 min. Liquid chromatography tandem-mass spectrometry (LC-MS/MS) was employed to confirm the performance of the strip. In addition, a novel pretreatment was established and compared with conventional pretreatment. Without the removal of organic solvents, the novel pretreatment method reduced the sample pretreatment time (more than 10 min). The vLOD of the optimized GICA was also 2 ng·g^-1 when applying the novel pretreatment method. In conclusion, the proposed PEF-GICA could detect samples containing PEF rapidly and accurately, and the novel pretreatment method saved the time of sample pretreatment and improved the efficiency of detection.