| 翻译控制肿瘤蛋白TPT1的原核表达、纯化、抗体制备和初步应用 |
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| 引用本文： | 赵一璠,李海芳,汤石明,王萌,吴海东,刘民,李欣,汤华.翻译控制肿瘤蛋白TPT1的原核表达、纯化、抗体制备和初步应用[J].细胞与分子免疫学杂志,2010,26(3). |
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| 作者姓名： | 赵一璠 李海芳 汤石明 王萌 吴海东 刘民 李欣 汤华 |
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| 作者单位： | 天津医科大学基础医学院生命科学中心实验室,天津,300070 |
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| 基金项目： | 国家自然科学基金资助项目，天津市自然科学基金重点项目，国家高技术研究发展计划(863)资助项目 |
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| 摘 要： | 目的:原核表达并纯化翻译控制蛋白TPT1,免疫日本大耳白兔制备其抗体。方法:构建原核表达质粒pRSE-TA2-TPT1,转化大肠杆菌BL21(DE3),TPT1蛋白经异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达。融合蛋白通过Ni-NTA树脂亲和纯化后免疫兔子制备抗体血清。以间接ELISA法检测抗体效价,Western blot和免疫荧光染色鉴定抗体特异性。结果:在大肠埃希菌中诱导出高水平表达的TPT1融合蛋白,经亲和树脂纯化后免疫大白兔,获得了高特异性的抗TPT1抗血清。结论:成功构建原核表达质粒pRSETA2-TPT1,表达并纯化TPT1蛋白,制备出高滴度、高特异性的多克隆抗体.为进一步研究TPT1在肿瘤等疾病发生、发展过程及治疗中的作用奠定基础。
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| 关 键 词： | 原核表达 纯化 抗体 肿瘤 |
Expression,purification,antibody preparation and identification of tumor protein translationally-controlled 1(TPT1) |
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| ZHAO Yi-fan,LI Hai-fang,TANG Shi-ming,WANG Meng,WU Hai-dong,LIU Min,LI Xin,TANG Hua.Expression,purification,antibody preparation and identification of tumor protein translationally-controlled 1(TPT1)[J].Journal of Cellular and Molecular Immunology,2010,26(3). |
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| Authors: | ZHAO Yi-fan LI Hai-fang TANG Shi-ming WANG Meng WU Hai-dong LIU Min LI Xin TANG Hua |
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| Institution: | ZHAO Yi-fan,LI Hai-fang,TANG Shi-ming,WANG Meng,WU Hai-dong,LIU Min,LI Xin,TANG Hua Tianjin Life Science Research Center,Basic Medical School,Tianjin Medical University,Tianjin 300070,China |
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| Abstract: | AIM:To express and purify the fusion protein of TPT1(tumor protein translationally-controlled 1) in prokaryocytes and to prepare rabbit anti-TPT1 antibody.METHODS:The expression vector pRSETA2-TPT1 was reconstructed and transformed into BL21(DE3).TPT1 fusion protein was induced by IPTG and the TPT1 fusion protein purified by Ni-NTA His Bind resin was used to immunize the rabbit.The titer of the polyclonal antibody was detected by ELISA and its specificity was analyzed by Western blot and IF(immunofluorescen... |
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| Keywords: | TPT1 TPT1 prokaryotic expression purification antibody tumor |
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