Competitive immunoassay for simultaneous detection of imidacloprid and thiacloprid by upconversion nanoparticles and magnetic nanoparticles

A rapid and sensitive immunoassay for the simultaneous detection of imidacloprid and thiacloprid was developed by using magnetic nanoparticles (MNPs) and upconversion nanoparticles (UCNPs). The UCNPs of NaYF₄:Yb, Er and NaYF₄:Yb, Tm were synthesized and conjugated with anti-imidacloprid monoclonal antibody (mAb) and anti-thiacloprid mAb as signal labels, while the MNPs were conjugated with antigens of thiacloprid and imidacloprid as separation elements. The fluorescence intensities of Yb/Er- and Yb/Tm-doped UCNPs were detected simultaneously in 544 nm and 477 nm under the excitation of NIR light (980 nm). The amounts of mAb-conjugated UCNPs that were separated by antigen-conjugated MNPs were determined based on competitive immunoassays. Under the optimal conditions, the 50% inhibiting concentration (IC₅₀) and limit of detection (LOD, IC₁₀) were 5.80 and 0.32 ng/mL for imidacloprid and 6.45 and 0.61 ng/mL for thiacloprid, respectively. The immunoassay exhibited negligible cross-reactivity with analogs of imidacloprid and thiacloprid except imidaclothiz (86.2%). The average recoveries of imidacloprid and thiacloprid in environmental and agricultural samples, including paddy water, soil, pears, oranges, cucumbers, and wheat, ranged from 78.4 to 105.9% with relative standard deviations (RSDs) of 2.1-11.9% for imidacloprid and ranged from 82.5 to 102.3% with RSDs of 1.0-16.5% for thiacloprid. In addition, the results of the immunoassay correlated well with high-performance liquid chromatography for the detection of the authentic samples.