Selection of a Single Chain Variable Fragment Antibody against Ivermectin from a Phage Displayed Library
Selection of a Single Chain Variable Fragment Antibody against Ivermectin from a Phage Displayed Library
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从噬菌体展示文库中选择抗伊维菌素单链可变片段抗体
DOI：
10.1021/jf904562x
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发表时间：
2010-05-12
影响因子：
6.1
通讯作者：
Liang, Jiansheng
中科院分区：
文献类型：
--
作者：
Wen, Shuang;Zhang, Xiao;Liang, Jiansheng
单链可变区抗体(ScFv)在农药、兽药和其他残留的免疫学检测中具有很大的潜力。本研究利用人合成的大容量噬菌体展示文库，通过消减淘选法筛选抗伊维菌素(IVM)的单链抗体。经过4轮淘洗，获得了7个能与IVM结合的单克隆噬菌体颗粒。在大肠杆菌HB2151中表达了一种新的抗IVM单链抗体hsIVM8，并用镍金属离子亲和层析对其进行纯化。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法(SOS-PAGE)表明，hsIVM8的相对分子质量为28 kDa。将纯化的hsIVM8用于建立检测IVM的竞争性间接酶联免疫吸附试验(CI-ELISA法)。该方法的线性范围约为0.1~5mU g/m L，对照的半数抑制浓度(IC50)为4.11 m u g/m L。这些结果表明，hsIVM8在检测农业和环境基质中的IVM方面具有良好的应用前景。
Single chain variable fragment antibodies (scFvs) have considerable potential in immunological detection of pesticides, veterinary and other residues. In this study, a large human synthetic phage displayed library was used to select scFvs against ivermectin (IVM) by subtractive panning. After four rounds of panning, seven monoclonal phage particles capable of binding with the IVM were obtained. One of the identified novel anti-IVM scFv, hsIVM8, was expressed in Escherichia coli HB2151 and purified with Ni metal ion affinity chromatography. Sodium dodecyl sulfate polyacrylamine gel electrophoresis (SOS PAGE) indicated that the relative molecular mass of hsIVM8 was estimated at 28 kDa. The purified hsIVM8 was used to develop a competitive indirect enzyme-linked immunosorbent assay (ci-ELISA) for IVM. The linear range of detection for standards in this ci-ELISA was approximately 0.1-5 mu g/mL and 50% inhibition of control (IC50) was 4.11 mu g/mL for IVM. These results showed promising applications of hsIVM8 for detecting IVM in agricultural and environmental matrices.