将异丙甲草胺与3-巯基丙酸反应合成出带羧基的异丙甲草胺半抗原MMPA;用活泼酯法将半抗原与载体牛血清白蛋白(BSA)和卵清白蛋白(OVA)分别偶联,制备免疫抗原MMPA-BSA和包被抗原MMPA-OVA,紫外光谱鉴定后用MMPA-BSA免疫新西兰大白兔,成功制备出抗异丙甲草胺多克隆抗体,并建立异丙甲草胺的间接竞争酶联免疫吸附分析(icELISA)法。该方法半抑制浓度(IC50)为34.3ng/mL,检测限(IC10)为6.3ng/mL,检测范围(IC20~IC80)为12.3~99.2ng/mL。该抗体对其他结构类似物无明显交叉反应。将该方法应用于环境水样中异丙甲草胺检测,回收率在89.5%~107.9%之间,相对标准偏差9.2%~14.5%。所建立的异丙甲草胺免疫分析方法特异性强、准确度高、灵敏度高,可用于环境水样中异丙甲草胺的快速检测。
Using one step reaction,metolachlor was modified with 3-marcapropanoic acid(3-MPA) to obtain the hapten 3-(2-((2-ethyl-6-methylphenyl)(1-methoxypropan-2-yl)amino)-2-oxoethylthio)propanoic acid(MMPA),which was then coupled to bovine serum albumin(BSA) as immunogen(MMPA-BSA) and ovalbumin(OVA) as coating antigen(MMPAOVA) using active ester method,respectively.New Zealand rabbits were immunized with MMPA-BSA.Based on the obtained polyclonal antibody,an indirect competitive ELSIA(icELISA) was developed successfully for the detection of metolachlor and demonstrated excellent performance.The 50% inhibition concentration(IC50) was 34.3 ng/mL and the detectable range(IC20–IC80) was 12.3–99.2 ng/mL,while the limit of detection(LOD,IC10) was 6.3 ng/mL.The antibody showed low cross-reactivity(10.9%) towards S-(-)-metolachlor.However,no significant cross-reactivity towards other structurally related compounds was found.The recoveries of spiked samples were in the range of 89.5%–107.9% with RSD ranging from 9.2% to 14.5%.Our results indicated that the proposed icELISA method is highly specific,sensitive,accurate and suitable for the rapid detection of herbicide metolachlor in water samples.