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分析化学 2012

呋喃唑酮代谢物单克隆抗体制备及酶联免疫吸附分析方法

DOI: 10.3724/SP.J.1096.2012.10409, PP. 745-751

任海涛,沈玉栋,徐振林,杨金易,雷红涛,肖治理,王弘,孙远明

Keywords: 呋喃唑酮,单克隆抗体,酶联免疫法,异源包被

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Abstract:

本研究针对呋喃唑酮代谢物(AOZ),设计合成了系列半抗原,进一步通过偶联牛血清白蛋白(BSA)免疫Balb/c小鼠、细胞融合、筛选和亚克隆等过程成功获得了源于新颖半抗原H3的具有高亲和力(亲和力常数6.68×1010L/mol)和高特异性(与其它功能类似物交叉反应小于0.1%)抗AOZ单克隆抗体。同时,基于设计合成的系列同/异源半抗原/包被抗原,考察了不同结构包被原对ELISA方法灵敏度的影响。另外,采用最佳的特征结构异源包被原H5-OVA,建立了以对硝基苯甲醛(p-NP)为衍生剂的AOZ间接竞争ELISA(icELISA)和直接竞争ELISA(dcELISA)检测方法。结果表明:icELISA模式的AOZ检测IC50为0.503μg/L,定量检测线性范围(IC20~IC80)为0.06~14.0μg/L,检出限(IC10)达0.017μg/L;dcELISA模式的AOZ检测IC50为1.19μg/L,定量检测线性范围为0.14~23.6μg/L,检出限为0.056μg/L。两种方法对AOZ的检测灵敏度和定量线性范围均达到相关检测限量要求,可满足不同需求的实际样品检测。

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