Chemiluminescent Enzyme Immunoassay For Rapid Detection Of Three α-cyano Pyrethroid Residues In Agricultural Products

Synthetic pyrethroids are widely used in domestic, public health, agricultural, forestry, and veterinary applications for their remarkably high insecticidal activity and low mammalian toxicity compared with organochlorinated and organophosphate compounds. The α-cyano pyrethroids such as deltamethrin, cypermethrin,λ-cyhalothrin, fenpropathrin and fenvalerate, are quickly becoming the dominant. Although pyrethroids are thought to be safe for humans, due to the extensive human use and release of synthetic pyrethroids, their toxic effects on ecological, environmental, and human health have become increasingly concerned. Therefore, it is necessary to determine α-cyano pyrethroid residues in the environment and agricultural products for regulatory and monitoring purposes.In this study, a new monoclonal antibody (mAb) was first produced based on a genetic hapten containing the common moiety of α-cyano pyrethroids but without one side chain of the cyclopropane ring. A comptitive indirect enzyme-linked immunosorbent assay (ELISA) was established to characterize the mAb, using the antigen-coated format. Afterward, assay condition was optimized for pH and ionic strength of assay buffer, organic solvent concentration and reaction time. The optimal ELISA was broad-specific to three a-cyano pyrethroids, and the half-maximal inhibition concentration (IC50) of fenpropathrin, delthamethrin and λ-cyhalothrin were 2.6,8.2 and 31.1 ng mL-1. The cross-reactivity to other pyrethroids was not obvious, except 1.7% for cypermethrin.To further improve the assay performances, an indirect competitive chemiluminescent enzyme immunoassay (CLEIA) was employed by introducing an enhanced chemiluminescent (ECL) reaction as the end-point detection system. Under the optimum conditions, the CLEIA showed IC50 values of 1.9,3.4 and 4.3 ng mL-1 for fenpropathrin, delthamethrin and λ-cyhalothrin. The cross-reactivity to other pyrethroids was also not obvious, except 5% for cypermethrin. Compared with the colorimetric ELISA, the ability of the chemiluminescent reagents to detect lower concentrations of horseradish peroxidase did allow to decrease the consumption of antibody and antigen (antibody concentration inCLEIA was two-fold less than that used in ELISA, and antigen concentration was four-fold less). Additionally, the CLEIA presented other advantages over the ELISA, such as better assay sensitivity and less reaction time.Moreover, the immuoassay application was performed to determination fenpropathrin, deltamethrin and λ-cyhalothrin tap in water and agriculture samples (orange, eggplant, and cowpea samples), using recovery tests at different spiked levels. Matrix interferences on the standard calibration curves were also investigated to determine suitable dilution times for sample extracts. It was found that CLEIA was more tolerant to sample matrices than ELISA, since less dilution times were needed in CLEIA for both water and agriculture samples. Overall, recovery results obtained from CLEIA was correlated well with those from the instrumental method of gas chromatography equipped with an electron capture detector (GC-ECD), both in terms of accuracy and precision.To the best of our knowledge, this is the first study on CLEIA method for the detection of a-cyano pyrethroids, which is suitable as a rapid screening tool for the control of pesticide contamination in agricultural products.