| 犬γ干扰素多克隆抗体的制备及生物活性检测 |
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| 引用本文： | 李一经,吴昱,唐丽杰,葛俊伟,乔薪瑗,崔文,姜艳平.犬γ干扰素多克隆抗体的制备及生物活性检测[J].东北农业大学学报,2012(12):73-77. |
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| 作者姓名： | 李一经 吴昱 唐丽杰 葛俊伟 乔薪瑗 崔文 姜艳平 |
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| 作者单位： | 东北农业大学动物医学学院 |
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| 基金项目： | 黑龙江省高校科技创新基金(2011TD001) |
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| 摘 要： | 将犬γ干扰素基因和表达载体pET30a双酶切后构建了重组质粒pET30a-CaIFN-γ,并转化到E.coliRosetta感受态中进行诱导表达。SDS-PAGE和Western blot分析显示,重组蛋白约23 ku并主要以包涵体形式存在。包涵体通过切胶纯化后肌注家兔,制备出CaIFN-γ的抗血清,测得血清效价为1:12 800。对复性后的包涵体通过细胞试验进行抗病毒活性检测,表明重组犬γ干扰素在终浓度为0.09 mg·mL-1时具有一定的抗病毒活性。本试验成功的原核表达、纯化了犬γ干扰素蛋白,制备CaIFN-γ的抗血清,为CaIFN-γ蛋白生物学特深入研究和生产应用奠定基础。
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| 关 键 词： | 犬IFN-γ 原核表达 抗病毒活性 多克隆抗体 |
Preparation of rabbit antisera against CaIFN-γ and evaluation of CaIFN-γ bioactivity |
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| LI Yijing,WU Yu,TANG Lijie,GE Junwei,QIAO Xinyuan,CUI Wen,JIANG Yanping.Preparation of rabbit antisera against CaIFN-γ and evaluation of CaIFN-γ bioactivity[J].Journal of Northeast Agricultural University,2012(12):73-77. |
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| Authors: | LI Yijing WU Yu TANG Lijie GE Junwei QIAO Xinyuan CUI Wen JIANG Yanping |
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| Institution: | (School of Veterinary Medicines,Northeast Agricultural University,Harbin 150030,China) |
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| Abstract: | The canine IFN-γ gene was digested and was inserted into plasmid pET30a to construct recombinant plasmid pET30a-CaIFN-γ,then the recombinant plasmid was transformed into E.coil Rosetta.The recombinant protein was expressed after inducing.SDS-PAGE and Western-blot results showed that the recombinant protein was about 23 ku and mainly in the form of inclusion bodies.CaIFN-γ antiserum was generated by immunizing the rabbit with the purified recombinant CaIFN-γ protein by gel isolation.ELISA result showed that the serum titer was 1∶12 800.The biological activity of renaturated protein was checked by cell experiment.Cell experiment result showed that CaIFN-γ with the terminal concentration of 0.09 mg.mL-1 possesses characteristic of antivirus.The experiment successfully expressed and purified the CaIFN-γ protein and obtained the CaIFN-γ antiserum,and provided a reliable basis for the further study on CaIFN-γ protein biological characteristics and its application in the productive practice. |
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| Keywords: | CaIFN-γ prokaryotic expression antiviral activity polyclonal antibody |
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