A Sensitive Lateral Flow Immunoassay Relying on Time-Resolved Fluorescent Microspheres Immune Probe for Determination of Ceftiofur and Its Metabolite

Ceftiofur (CEF) is an antimicrobial agent with high efficiency and low toxicity, desfuroylceftiofur is its main metabolite, but they are also have potential harm to human health. In this study, ceftiofur was combined with carrier proteins to get artificial antigens. A specific antibody (pAb) against CEF and desfuroylceftiofur was prepared. A sensitive and rapid paper-based sensor relying on time-resolved fluorescent microspheres (TRFMs) immune probes was developed, which were time-resolved fluorescent immunochromatographic strips (TRFMsLFIA). The concentrations of T line and C line, activated pH, antibody volume and probe volume were optimized. Quantitative limits of detection (qLODs) of TRFMs-LFIA for CEF and desfuroylceftiofur were 0.97 ng/mL and 0.41 ng/mL, respectively. And 50 % inhibiting concentrations (IC50) were 12.92 ng/mL and 12.58 ng/mL, respectively. Pretreatment procedures of real samples were simple and rapid. Detection time of TRFMs-LFIA strip was 15 min. Qualitative analysis of CEF and desfuroylceftiofur was achieved under a UV light, quantitative analysis was implemented with a fluorescent immunoassay analyzer. The average recovery rates ranged from 91.4 % to 107.7 % and corresponding coefficients of variation (CV) was 1.5%-9.7 %. Concentration levels of artificially-spiked samples were measured by TRFMs-LFIA and compared with detection results of High performance liquid chromatography (HPLC), which showed a good accordance. These results indicated that the proposed assay can provide an effective strategy for on-site detection of CEF and desfuroylceftiofur simultaneously.