Development of an ic-ELISA and a TRFICA for the detection of fipronil with a new hapten design

Based on the new hapten of fipronil, an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and time-resolved fluorescent immunochromatographic assay (TRFICA) for the determination of fipronil were developed. The sensitivity (IC₅₀) of the ic-ELISA method was 2.33 ± 0.14 ng mL⁻¹ (n = 3), and the limit of detection (LOD) (IC₂₀) was 0.27 ± 0.02 ng mL⁻¹ (n = 3). The cross-reaction rates of the three fipronil metabolites were all higher than 66.7%, and those of the three analogs were no more than 0.05%. The TRFICA was optimized with a handheld reader. The LOD was 0.862 ng mL⁻¹ (n = 3). In this study, the pretreatment methods of samples such as Chinese cabbage, spinach, apple and egg, were explored. Methanol and acetonitrile were used to extract different food samples and the diluents were optimized. Three concentration levels in the spiked food samples were detected by the two methods. The range of recovery was 88.65–116.32% for ic-ELISA and 95.48–112.61% for TRFICA. In this study, the accuracy of the two methods was also judged by testing commercial quality control samples. The results showed that the accuracy was 83.75%–113.33% for ic-ELISA and 98.33–107.13% for TRFICA.