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Production and Identification of Broad Spectrum Monoclonal Antibody against a Group of Pyrethroid Insecticides

文献类型： 外文期刊

作者： Wen Meng-Tang 1 ; Liu Yuan 1 ; Yan Shuai 1 ; Zhang Xiao 1 ; Wang Heng 1 ; Liu Xian-Jin 1 ;

作者机构： 1.Jiangsu Acad Agr Sci, Inst Food Safety, Nanjing 210014, Jiangsu, Peoples R China

2.Northwest Agr & Forestry Univ, Coll Food Sci & Engn, Yangling 712100, Peoples R China

关键词： Pyrethroid pesticides;Broad specificity;Monoclonal antibody;Enzyme-linked immunosorbent assay

期刊名称：CHINESE JOURNAL OF ANALYTICAL CHEMISTRY （ 影响因子：1.134； 五年影响因子：0.909 ）

ISSN： 0253-3820

年卷期： 2014 年 42 卷 9 期

页码：

收录情况： SCI

摘要： The objective of this study is to generate broad spectrum monoclonal antibody(mAb) against a group of pyrethroid insecticides and to identify its immunological characteristics. The generic hapten 3-phenoxy-benzoic acid (PBA) was conjugated to carrier protein BSA by activated ester method. Balb/c mice were immunized with PBA-BSA. The titer of polyclonal antibody (pAb) was detected by indirect enzyme-linked immunosorbent assay (ELISA) after five times immunization. The mouse with high titer and sensitivity was selected for cell fusing. The splenocytes of immunized mice were fused with Sp2/0 cells and the cultural supernatants of hybridoma cells were screened by indirect non-competitive ELISA based on the coating antigen PBA-ovoalbumin (PBA-OVA). High-sensitivity and high-specificity mAb was prepared after subcloning using limiting dilution method. Purified mAb was obtained after purified by saturated ammonia sulfate precipitation and protein G affinity column. The immunological characteristics of mAb such as titer, antibody subtypes, affinity constant and the sensitivity to pyrethroid insecticides were characterized by indirect ELISA; The results of UV spectroscopy and SDS-PAGE showed that PBA-BSA artificial antigen was synthesized successfully. A hybridoma cell line (4H11) secreting anti-pyrethroid mAb was established. The titre of ascites was up to 1: 6. 5x10(6), and the mAb was IgG1 subtype. The affinity constant of the mAb to PBA was about 2. 5x10(7) L/mol, with a IC50 value of 208. 83 mu g/L and a detection limit of 21. 23 mu g/L to PBA. Simultaneously, beta-cypermethrin, flucythrinate, cypermethrin and fenvalerate were sensitively recognized by the mAb with the IC50 of 1. 01, 2. 15, 3. 16 and 3. 67 mu g/L, respectively.

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