菊酯类农药广谱型免疫层析试纸条的研究及应用

Lateral Flow Immunoassay for Simultaneous Determination of Four Kinds of Pyrethroid Pesticides in Vegetables and Fruits

建立了菊酯类农药广谱型免疫层析检测方法,可同时检测水果、蔬菜中12种甲氰菊酯、溴氰菊酯、氯氰菊酯、三氟氯氰菊酯农药残留。以粒径20 nm的胶体金标记羊抗小鼠IgG抗体于金标垫上,分别固定包被原（检测线,T线）、兔抗山羊IgG抗体（质控线,C线）于硝酸纤维素膜（NC膜）上,与吸水垫及聚氯乙烯（PVC）底板组合成层析试纸条。10 g样品经乙腈提取,PBS稀释4倍,取100μL与菊酯类农药的单克隆抗体混合后,直接用于试纸条检测。结果表明,试纸条对甲氰菊酯、溴氰菊酯、氯氰菊酯、三氟氯氰菊酯农药的裸眼观察检测灵敏度为0.5,5.0,5.0和5.0μg/mL,检测时长为8~10 min,采用QuEChERS方法,方法检测灵敏度可提高16倍。对蔬菜和水果的方法验证表明,除辣椒基质干扰呈假阳性外,其它样本的检测结果均与GC方法结果一致。试纸条采用金标羊抗小鼠IgG二抗的方法,使检测结果的重现性更好、更稳定,且抗基质干扰能力强,为菊酯类农药的累积毒性检测提供了新方法。

A monoclonal antibody based lateral flow immunoassay was developed for the simultaneous determination of four pyrethroid pesticides in vegetables and fruits, including fenpropathrin, cypermethrin, deltamethrin, and cyhalothrin. The colloidal gold particles （20-nm） were prepared by the method of sodium citrate reduction and conjugated with goat anti-mice IgG. Antigen and rabbit anti-goat IgG were immobilized on the nitrocellulose membrane as the test line （ T line） and the control line （ C line） , separately. By sticking on PVC board, a gold immunochromatographic strip was assembled. About 10 g of samples was extracted by acetonitrile, and then diluted four times with PBS buffer. Anti-pyrethroid McAb was added in 100 μL of this extract solution for the antibody-antigen binding, thereafter, the mixture was ready for the strip testing. The results revealed that the test could be done within 10 min, and the cutoff values reached 0. 5, 5. 0, 5. 0, 5. 0 μg/mL for fenpropathrin, cypermethrin, deltamethrin, cyhalothrin, respectively. If the extraction going through QuEChERS method, the detection limit of method could be improved 16 times. Thereafter, the lateral flow immunoassay was employed for the verification test with 13 types of vegetables and fruits, except that the result of chillies presented the false positive result, the other results obtained higher consistency with the results of GC test. This new format of test strip using colloidal gold labeled goat anti-mice IgG as tracer, which could give more stable and repeatable results than classic test strip, would provide a new method for the simultaneous determination of four pyrethroid pesticides in vegetables and fruits.