抗幽门螺杆菌细胞空泡毒素纳米抗体的制备及鉴定  

Preparation and Identification of Anti-VacA of Helicobacter pylori Nanobody

作　　者：刘晓芳 刘琼 黄云祥 钟引凤 何庆华 李燕萍 涂追 付金衡 LIU Xiaofang;LIU Qiong;HUANG Yunxiang;ZHONG Yinfeng;HE Qinghua;LI Yanping;TU Zhui;FU Jinheng(Institute of Life Sciences,Nanchang University,Jiangxi Nanchang 330031;Sino-Germany Joint Research Institute,Nanchang University,Jiangxi Nanchang 330047;State Key Laboratory of Food Science and Technology,Nanchang University,Jiangxi Nanchang 330047;Department of Medical Microbiology,School of Medicine,Nanchang University,Jiangxi Nanchang 330031)

机构地区：[1]南昌大学生命科学学院,江西南昌330031 [2]中德联合研究院,江西南昌330047 [3]食品科学与技术国家重点实验室,江西南昌330047 [4]南昌大学基础医学院,江西南昌330031

出　　处：《生物化工》2021年第1期11-15,22,共6页Biological Chemical Engineering

摘　　要：目的:获取抗幽门螺杆菌(Helicobacter pylori,Hp)细胞空泡毒素纳米抗体并鉴定其结合和抑制幽门螺杆菌的能力。方法:用灭活的幽门螺杆菌免疫羊驼,以重组蛋白VacA为靶点,从噬菌体展示文库中筛选与幽门螺杆菌特异性结合的纳米抗体,间接phage-ELISA鉴定结合重组蛋白VacA能力并诱导其进行原核表达,鉴定其与Hp的结合力。结果:获得5×107库容的噬菌体文库,将筛选的OD450值较高的纳米抗体HV2-36表达,经鉴定为可溶性蛋白且表达量为102.8 mg/L;所制备纳米抗体能特异性结合Hp菌体蛋白,并对Hp具有体外抑制作用。结论:本研究成功获得抗幽门螺杆菌VacA的纳米抗体。Objective:Obtained anti-Helicobacter pylori cell vacuolating toxin nanobody and identified the ability to bind and inhibit Helicobacter pylori.Methods:The alpaca was immunized with the lysate of inactivated Hp.Using the recombinant protein VacA as target molecule,nanobodies that can specifically recognize recombinant protein VacA were screened from phage displayed immune libraries.Indirect phage-ELISA identifies the ability to bind recombinant protein VacA and induces the expression of prokaryotic by IPTG,and indirect ELISA identifies the binding ability to Hp.Result:A 5×107 phage display library is obtained.The screened nanobodies HV2-36 with high OD450 in phage-ELISA was recombinantly expressed.SDS-PAGE analysis showed that the production was soluble and the production yield was 102.8 mg/L.ELISA showed that production which was recombinantly expressed could bind to the lysate of Hp.The production could inhibit Hp in vitro.Conclusion:nanobodies against VacA of Helicobacter pylori are successfully obtained.

关 键 词：VACA 幽门螺杆菌 纳米抗体 phage-ELISA 

分 类 号：Q78[生物学—分子生物学]