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| 人源抗HBsAg抗体Fab段在酵母中的表达 | |
| 引用本文： | 宋宏彬,唐晓敏,胡小华,李申龙,刘雪林,邱立建,李锋. 人源抗HBsAg抗体Fab段在酵母中的表达[J]. 中国公共卫生, 2004, 20(10): 1186-1188 |
| 作者姓名： | 宋宏彬 唐晓敏 胡小华 李申龙 刘雪林 邱立建 李锋 |
| 作者单位： | 军事医学科学院疾病预防控制中心,北京,100039 |
| 基金项目： | 北京市自然科学基金资助 (70 0 2 0 33) |
| 摘 要： | 目的探索抗体在毕赤酵母中表达的可能性.方法首先构建既可在体外构建多拷贝、又可于体内筛选多拷贝的分泌型酵母表达载体-pD89,在此基础上插入抗乙型肝炎病毒表面抗原(HBsAg)人源抗体Fab段的基因,电转化后,经G418筛选,得到阳性重组克隆.结果聚丙烯凝脉电泳(SDS-PAGE)检测证明,甲醇诱导后,阳性重组克隆可分泌表达分子量约50 KD的蛋白,免疫印迹实验证实,该蛋白为人源抗体Fab段.结论ELISA结果显示,表达的Fab段能够与HBsAg结合. |
| 关 键 词： | 基因工程抗体 表达 毕赤酵母 HBsAg |
| 文章编号： | 1001-0580(2004)10-1186-03 |
| 修稿时间： | 2004-01-23 |
Functional expression of human antibody fab fragment against hepatitis virus B surface antigen in Pichia pastoris |
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| SONG Hong-bi n,TANG Xia o-min,HU Xiao-hua,et al.. Functional expression of human antibody fab fragment against hepatitis virus B surface antigen in Pichia pastoris[J]. Chinese Journal of Public Health, 2004, 20(10): 1186-1188 | |
| Authors: | SONG Hong-bi n TANG Xia o-min HU Xiao-hua et al. |
| Affiliation: | SONG Hong-bi n,TANG Xia o-min,HU Xiao-hua,et al.Center for Disease Control and Prevention,Academy of M ilitary Medical Sciences |
| Abstract: | Objective To explor e the possibility of antibody production in Pichia pastoris.Methods A new type of Pichia pastoris expression vector,pD89,was constructed,which f aciliated to construct multiple copy inserts in vitro and screen multiple copy inserts in vivo .The Light chain and Fd fragment gene of the human antibody against Hepatitis Vi rus B Surface Antigen(HBsAg) were then inserted to pD89 under the control of AO X 1 promoter with a secretion signal in different expression cassette.After electr oporation and screening with G418,7 Mut clones with different G418 resistance were selected for expression.Results Results of SDS-PAGE showed that 3 clones expresse d a 50 KD protein.The protein was further confirmed to be a human Fab fragment b y western blot. Conclusion Binding activity to HBsAg was characterized by ELISA. |
| Keywords: | engineering antibody expression Pichia pasto ris HBsAg |
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