Chen Xiao-Hua, Cai Guo-Ping
Department of Biological Science and Biotechnology, Tsinghua University, Beijing, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2007 Oct;23(10):950-2.
To express the recombinant proteins of human fibrogenic factors and prepare the polyclonal antibodies of rabbit-anti-human fibrogenic factors.
The human fibrogenic factors were variant splicing forms of one gene. Their consensus sequence (ChS) was cloned into plasmid pQE30 to construct recombinant prokaryotic expression system. The recombinant expression vectors were transformed into E.coli M15, and then the expression vectors were induced by IPTG and many insoluble inclusion body proteins was obtained. The obtained proteins were used to immunize rabbits to obtain polyclonal antiserum with high titer and specificity. The eukaryotic expression of the recombinants of human fibrogenic factors was detected by Western blot.
The insoluble inclusion body proteins were also used to prepare polyclonal antibodies with high titer and specificity.
The expression of the recombinant proteins of human fibrogenic factors and the preparation of polyclonal antiserum provide a useful tool for further study of the expression and function of human fibrogenic factors.
表达人致纤维化因子的重组蛋白并制备兔抗人致纤维化因子多克隆抗体。
人致纤维化因子为一个基因的可变剪接形式。将其共有序列(ChS)克隆至质粒pQE30构建重组原核表达系统。将重组表达载体转化至大肠杆菌M15,然后用异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达载体,获得大量不溶性包涵体蛋白。用获得的蛋白免疫兔子,得到高滴度、特异性强的多克隆抗血清。通过蛋白质印迹法检测人致纤维化因子重组体的真核表达。
不溶性包涵体蛋白也可用于制备高滴度、特异性强的多克隆抗体。
人致纤维化因子重组蛋白的表达及多克隆抗血清的制备为进一步研究人致纤维化因子的表达及功能提供了有用工具。