| An indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed for the detection of nitenpyram.The hapten was synthesized successfully and conjugated to BSA and OVA to prepare immunogen and coating antigen by the active ester method, respectively. New Zealand white rabbits were immunized and the polyclonal antiserum with high specificity for nitenpyram was obtained.The indirect competitive ELISA was developed and the best results were obtained when the coating antigen concentration was 0.05μg/well and the antibody dilution was 1:2000. The blocking solution and assay buffer were selected as 0.5% skim milk power/PBS and 0.05 mol L-1 PBS. The half-maximum inhibition concentration (IC50) and limit of detection (LOD, calculated as IC15 value) of the ELISA for nitenpyram were 2.18± 0.22 ng mL-1 and 0.20±0.02 ng mL-1, respectively. The assay showed low cross-reactivity with imidacloprid (1.49%), acetamiprid (0.27%), thiacloprid (0.28%) and other pesticides (<0.01%), which indicated that the assay had high specificity. The assay was used to detect four spiked water samples (purified, tap, well and river water), four spiked fruit juice samples (tomato, apple, orange and grape juice) and three spiked vegetable samples (tomato, cucumber and cabbage). The recoveries were ranged from 62.8% to 116.5% and the coefficients of variation of intra- and inter-assays were less than 10%. There was a strong correlation (R2=0.9986) between data obtained using ELISA and gas chromatography (GC), indicating the good performance of this ELISA.The ELISA allow for a rapid, sensitive, specific, and low-cost determination of nitenpyram residues in water, fruit juice and vegetable samples. |
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