Novel inner filter effect-based fluorescence immunoassay with gold nanoclusters for bromadiolone detection in human serum

Bromadiolone (BRD) is a widely used second-generation rodenticide and BRD poisoning in humans is relatively common. However, few rapid diagnostic methods for BRD poisoning are currently available. In this study, a highly sensitive and specific fluorescence immunoassay was developed for the rapid detection of BRD in human serum. Briefly, two novel BRD haptens were synthesized, a sensitive and specific mAb for BRD (15C1) was produced. We then incorporated mAb 15C1 as a detection reagent, and gold nanoclusters (AuNCs) as a fluorescent probe into an alkaline phosphatase-based competitive fluorescence immunoassay. The sensitivity of the immunoassay was significantly improved by introducing the fluorescence signal of the AuNCs through the facile inner-filter effect strategy. Our fluorescence immunoassay exhibited high specificity and achieved a limit of detection of 0.047 ng/mL for BRD, which was > 10 times better than the corresponding standard enzyme-linked immunosorbent assay using the same immune reagents. The recoveries in spiked human serum were 77.9%-85.6% with coefficients of variation (CVs) of 3.6%-9.8%. There was a good correlation (R-2 = 0.987) between the fluorescence immunoassay and ultra-performance liquid chromatography-tandem mass spectrometry. The fluorescence immunoassay could provide a valuable clinical tool for the rapid diagnosis of BRD poisoning.