内皮抑素的原核表达及多克隆抗体的制备
[Prokaryotic expression of endostatin and preparation of polyclonal antibody].
作者信息
Zhang Yi-ming, Wang Ruo-ning, Li Yan-li, Gu Dong-min, Cheng Zhi-xiang, Xiong Jun, De Wei, Chen Bing-ying
机构信息
Department of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing 210029, P. R. China.
出版信息
Ai Zheng. 2002 Sep;21(9):957-60.
BACKGROUND & OBJECTIVE: The aim of this study was to express and purify human endostatin and to prepare polyclonal antibody of mouse anti-human endostatin.
METHODS
The cDNA of endostatin was amplified by PCR, then recombined into prokaryotic expression vector and transformed into Escherichia coli BL21 for expression; the mice were immunized with purified products.
RESULTS
Prokaryotic expression vector pQE-30 of human endostatin was successfully constructed; the expression product was gained after pQE-30 was transferred into BL21. After purified by Ni affinity chromatography, the product was identified to be a single component by SDS-PAGE. Western blot analysis showed that high titer mouse anti-human endostatin polyclonal antibody was successfully prepared.
CONCLUSION
Highly purified expression product and prepared polyclonal antibody provide the necessary material for further study.
背景与目的
本研究旨在表达并纯化人内皮抑素，制备小鼠抗人内皮抑素多克隆抗体。
方法
通过PCR扩增内皮抑素的cDNA，然后重组到原核表达载体中并转化到大肠杆菌BL21中进行表达；用纯化产物免疫小鼠。
结果
成功构建了人内皮抑素的原核表达载体pQE-30；将pQE-30转入BL21后获得表达产物。经镍亲和层析纯化后，SDS-PAGE鉴定产物为单一成分。Western blot分析表明成功制备了高滴度的小鼠抗人内皮抑素多克隆抗体。
结论
高纯度的表达产物和制备的多克隆抗体为进一步研究提供了必要材料。
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