[1]杜晓明,李宁,宋春青,等.人源性促甲状腺激素受体抗体 Fab段抗体库的构建[J].国际内分泌代谢杂志,2015,(04):238-241.[doi:10.3760/cma.j.issn.1673-4157.2015.04.006]
　Du Xiaoming*,Li Ning,Song Chunqing,et al.Construction of Fab fragment library of human thyrotrophin receptor antibody[J].International Journal of Endocrinology and Metabolism,2015,(04):238-241.[doi:10.3760/cma.j.issn.1673-4157.2015.04.006]
点击复制

人源性促甲状腺激素受体抗体 Fab段抗体库的构建()
分享到：

《国际内分泌代谢杂志》[ISSN:1673-4157/CN:12-1383/R]

卷:
期数:
2015年04
页码:
238-241
栏目:
论著
出版日期:
2015-07-20

文章信息/Info

Title:
Construction of Fab fragment library of human thyrotrophin receptor antibody
作者:
杜晓明; 李宁; 宋春青; 方佩华
300140　天津市第四中心医院内分泌科（杜晓明，宋春青）；300052　天津医科大学总医院核医学科（李宁，方佩华）
Author(s):
Du Xiaoming*; Li Ning; Song Chunqing; Fang Peihua.
*Department of Endocrinology，Tianjin 4th Center Hospital，Tianjin 300140，China Corresponding author：Fang Peihua，Email：raysinomail@163.com
关键词:
噬菌体表面展示技术; 人源性单克隆抗体; 促甲状腺激素受体抗体 Fab
Keywords:
Phage display technology; Human monoclonal antibodies; Thyrotrophin receptor antibody Fab
DOI:
10.3760/cma.j.issn.1673-4157.2015.04.006
摘要:
目的　通过噬菌体表面展示技术，构建人源性促甲状腺激素受体抗体（TRAb）Fab片段抗体库。方法　从甲状腺功能亢进症患者外周血单个核细胞抽提总RNA，PCR法扩增免疫球蛋白分子轻链κ、λ基因及重链Fd基因。构建轻链文库及组合文库。一个随机的组合文库在噬菌体表面表达。结果　从外周血单个核细胞中抽提得到总RNA，并反转录获得cDNA文库。PCR法扩增了大小约为680 bp的轻链κ、λ基因及重链Fd基因，并构建了库容为1.32×105基因抗体库(轻链库)和库容为2.28×105 Fab抗体库（组合文库）。Fab组合文库转化到大肠杆菌XL1-Blue感受态细胞，在辅助噬菌体M13K07的辅助下，扩增得到噬菌体抗体库。结论　噬菌体表面展示技术可成功构建人源性TRAb Fab片段组合文库。
Abstract:
Objective　To construct a Fab fragment library of human thyrotrophin receptor antibody （TRAb） using phage display technology. Methods　Total RNA was isolated from peripheral blood mononuclear cells of patients with hyperthyroidism. Human immunoglobulin κ/λ light chain and heavy chains Fd genes were amplified by PCR. A light chain library and a combinatorial library were constructed respectively. A random combinatorial library was expressed on the surface of filamentous phage. Results　cDNA library was harvested successfully by reverse transcription technology from total RNA of peripheral blood mononuclear cells. Human immunoglobulin 680 bp κ/λ light chain and heavy chain Fd genes were obtained by PCR. The size of κ light chain library was 1.32×105.The actual diversity of Fab combinatorial library was 2.28×105.The Fab combinatorial library were transformed in E.coli XL1 Blue.The transformered cells were infected with M13K07 helper phage to amplify phage antibody Fabs. Conclusion　A human TRAb Fab fragment phage combinatorial library can be constructed by phage surface display technology.

参考文献/References:

[1]　Shukra AM，Sridevi NV，Dev Chandran，et al.Production of recombinant antibodies using bacteriophages[J].Eur J Microbiol Immunol（Bp），2014，4（2）：91-98. 　
[2]　陆慧琦，钱新宇，李爱民，等.人源性抗核抗体Fab片段的筛选及鉴定[J].第二军医大学学报，2008，29（1）：87-91. 　
[3]　Coronella JA，Tellman P，Truong TD，et al.Amplification of IgG VH and VL （Fab） from single human plasma cells and B cells[J].Nucleic Acids Res，2000，28（20）：E85. 　
[4]　Siddiqui MZ.Monoclonal antibodies as diagnostics；an appraisal[J].Indian J Pharm Sci，2010，72（1）：12-17. 　
[5]　Barbesino G，Tomer Y.Clinical review：clinical utility of TSH receptor antibodies[J].J Clin Endocrinol Metab，2013，98（6）：2247-2255. 　
[6]　Latif R，Morshed SA，Zaidi M，et al．The thyroid-stimulating hormone receptor：impact of thyroid-stimulating hormone and thyroid-stimulating hormone receptor antibodies on multimer-ization，cleavage，and signaling[J].Endocrinol Metab Clin North Am，2009，38（2）：319-341.

备注/Memo

备注/Memo:
基金项目：天津市卫生局科技重点项目(2013KR04) 　　通信作者：方佩华，Email：raysinomail@163.com
更新日期/Last Update: 2015-07-20