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| 抗H5N1禽流感病毒VHH抗体库的构建 | |
| 引用本文： | 严安,孙冰玉,夏立亮,吴标,包文静,车小燕,孙志伟,金维荣,赵国屏,熊慧,王颖.抗H5N1禽流感病毒VHH抗体库的构建[J].中国免疫学杂志,2011,27(1):44-49. |
| 作者姓名： | 严安 孙冰玉 夏立亮 吴标 包文静 车小燕 孙志伟 金维荣 赵国屏 熊慧 王颖 |
| 作者单位： | 1. 上海人类基因组研究中心,卫生部重点疾病与健康重点实验室,上海,201203 2. 上海市免疫学研究所,上海交通大学医学院,上海,200025 3. 南方医科大学,广州,510515 4. 北京军事医学科学院生物工程研究所,北京,100071 5. 上海华冠生物芯片有限公司,上海,201203 |
| 基金项目： | 国家863项目资助(2006AA02A252); 上海市教委第五期重点学科(J50207)资助 |
| 摘 要： | 目的:构建抗H5N1禽流感病毒的小羊驼免疫噬菌体重链可变区抗体库(VHH型抗体库),为抗H5N1的VHH抗体筛选奠定基础。方法:利用H5N1灭活疫苗免疫小羊驼,一定免疫时间后测定小羊驼外周血清中抗体中和活性,分离其外周淋巴细胞,利用RT-PCR方法得到VHH抗体片段。通过优化连接和电转化方法,将足量VHH片段与pCANTAB5E连接后电转入大肠杆菌TG1,获得VHH抗体基因库;检测基因库库容以及多样性,并采用血凝抑制试验对噬菌体抗体库进行初步功能性鉴定。结果:利用H5N1灭活疫苗免疫小羊驼四次后,其外周血清中抗体血清抑制效价可达1∶2 560,构建的VHH抗体基因库库容可达3×108,随机挑选14个抗体基因克隆进行测序鉴定,结果显示均为独立克隆,表明所建抗体库多样性好。上述基因库经辅助噬菌体拯救后,得到抗H5N1的噬菌体VHH型抗体初级库,对初级库进行血凝抑制试验,结果呈阳性,表明初级库中存在具有潜在中和活性的抗H5N1抗体。结论:结果表明,已成功构建抗H5N1禽流感病毒的小羊驼免疫噬菌体重链抗体库,为进一步筛选抗H5N1禽流感的重链抗体打下良好基础,并为H5N1的早期临床诊断和治疗提供新的手段。 |
| 关 键 词： | H5N1 噬菌体抗体库 重链可变区抗体 禽流感病毒 |
Construction of phage display VHH antibody library against avian H5N1 virus from alpaca |
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| YAN An,SUN Bing-Yu,XIA Li-Liang,WU Biao,BAO Wen-Jing,CHE Xiao-Yan,SUN Zhi-Wei,JIN Wei-Rong,ZHAO Guo-Ping,XIONG Hui,WANG Ying.Construction of phage display VHH antibody library against avian H5N1 virus from alpaca[J].Chinese Journal of Immunology,2011,27(1):44-49. | |
| Authors: | YAN An SUN Bing-Yu XIA Li-Liang WU Biao BAO Wen-Jing CHE Xiao-Yan SUN Zhi-Wei JIN Wei-Rong ZHAO Guo-Ping XIONG Hui WANG Ying |
| Institution: | YAN An,SUN Bing-Yu,XIA Li-Liang,WU Biao,BAO Wen-Jing,CHE Xiao-Yan,SUN Zhi-Wei,JIN Wei-Rong,ZHAO Guo-Ping,XIONG Hui,WANG Ying.Chinese National Human Genome Center at Shanghai,Shanghai 201203,China |
| Abstract: | Objective:To construct phage display variable domain of heavy chain antibody library(VHH antibody library) from alpaca immunized with inactivated H5N1 vaccine for the future screening of VHH antibodies against avian H5N1 influenza virus.Methods:The camelid species(alpaca,Lama pacos) was selected for immunization with inactivated H5N1 vaccine.Hemagglutinin inhibition(HI) assay of serum from immunized alpaca was performed against H5N1 avian influenza virus one week after the fourth immunization.Peripheral lym... |
| Keywords: | H5N1 |
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