| 溴氰菊酯酶联免疫吸附分析方法研究 |
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| 引用本文： | 张帮红,施海燕,王鸣华.溴氰菊酯酶联免疫吸附分析方法研究[J].农药学学报,2009,11(2):261-268. |
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| 作者姓名： | 张帮红 施海燕 王鸣华 |
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| 作者单位： | 1.南京农业大学 植物保护学院 农药科学系,农业部作物病虫害监测与防控重点开放实验室,江苏 南京 210095 |
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| 基金项目： | 国家高技术研究发展规划("863"计划)项目 |
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| 摘 要： | 建立了定量测定溴氰菊酯间接竞争酶联免疫吸附分析方法(ic-ELISA)。合成了半抗原1-羧基-(3'-苯氧基苯基)甲基-3-(2',2'-二溴乙烯基)-2,2-二甲基环丙基羧酸酯(Med)和N-2-(羧基丙基)氨基甲酰基-(3'-苯氧基苯基)甲基-3-(2',2'-二溴乙烯基)-2,2-二甲基环丙基羧酸酯(Di)。分别采用碳二亚胺法和混合酸酐法将半抗原与牛血清蛋白(BSA)和卵清蛋白(OVA)偶联制备了免疫原Di-BSA和包被原Di-OVA、Med-OVA。将制得的溴氰菊酯免疫原免疫动物获得多克隆抗体,经间接非竞争ELISA法测得其效价为2.5×105。通过对甲醇含量、离子强度、pH值等影响因素进行异源分析条件的优化,确立了溴氰菊酯间接竞争酶联免疫分析方法的最佳检测条件(30%甲醇、氯化钠0.4 mol/L、pH 7.5),并建立了标准竞争曲线。该方法的IC50值为0.55±0.05 mg/L,检测限(IC10)为3.76±0.35 μg/L,对大多数拟除虫菊酯无交叉反应。分别在自来水、河水和土壤样品中添加0.05 ~5.0 mg/L(或mg/kg)的溴氰菊酯,回收率分别为89.7% ~106.8%、82.4% ~101.7%、75.6% ~97.8%。
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| 关 键 词： | 溴氰菊酯 酶联免疫吸附分析 多克隆抗体 |
| 收稿时间： | 2008/11/7 0:00:00 |
Enzyme-linked Immunosorbent Assay for Deltamethrin |
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| ZHANG Bang-hong,SHI Hai-yan and WANG Ming-hua.Enzyme-linked Immunosorbent Assay for Deltamethrin[J].Chinese Journal of Pesticide Science,2009,11(2):261-268. |
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| Authors: | ZHANG Bang-hong SHI Hai-yan and WANG Ming-hua |
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| Institution: | 1.Department of Pesticide Science,College of Plant Protection,Nanjing Agricultural University,Key Laboratory ofMonitoring and Management of Crop Diseases and Pest Insects,Ministry of Agriculture,Nanjing 210095,China |
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| Abstract: | An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was developed for the quantitative detection of deltamethrin.The haptens of deltamethrin 1-carboxy-(3'-phenoxyphenyl)methyl-3-(2',2'-dibromoethenyl)-2,2-dimethylcyclopropanecarboxylate (Med) and N-2-(carboxy-propyl)carbamoyl-(3'-phenoxyphenyl)methyl-3-(2',2'-dibromoethenyl)-2,2-dimethylcyclopropane-carboxylate (Di) were synthesized.Di was conjugated with bovine serum albumin (BSA) as immunogen (Di-BSA) by carbodiimide method and coating antigens (Di-OVA,Med-OVA) of deltamethrin were prepared by conjugating these two haptens with ovalbumin (OVA) by mixed anhydride method.Polyclonal antibodies were obtained from rabbits immunized with immunogen and the titer of the freeze-dried powder (2.5×105) was determined by indirect non-competitive ELISA procedure.Standard competitive curve of the optimized ic-ELISA for deltamethrin was developed.Thus,the optimized assay conditions such as concentration of methanol (30%),inonic strength (NaCl 0.4 mol/L),pH value (7.5) were investigated and obtained by analysis of heterologous.The value of 50% inhibition of antibody binding (IC50) for deltamethrin was 0.55±0.05 mg/L,and the low detection limit (IC10) was 3.76±0.35 μg/L.This ELISA assay had no cross-reactions significantly with other major pyrethroids.Tap water,river water and soil samples were spiked with deltamethrin at 0.05~5.0 mg/L(or mg/kg),the average recoveries were in the range of 89.7% ~106.8%,82.4% ~101.7%,75.6% ~97.8% respectively. |
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| Keywords: | deltamethrin enzyme-linked immunosorbent assay polyclonal antibody |
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