丙型肝炎病毒非结构蛋白 3人源单链可变区抗体的筛选与鉴定(英文)  

Screening and identification of a humanized single chain variable region antibody for hepatitis C virus non-structural 3 protein

作　　者：成军[1] 钟彦伟[1] 刘妍[1] 董菁[1] 杨继珍[1] 张玲霞[1] 

机构地区：[1]解放军302医院传染病研究所基因治疗研究中心,北京100039

出　　处：《免疫学杂志》2000年第4期246-249,共4页Immunological Journal

基　　金：This work is supported by National Natural Science Foundation of China(3990 0 130 )

摘　　要：目的筛选、鉴定抗丙型肝炎病毒 (HCV)非结构蛋白 NS3的人单链可变区抗体 (Sc Fv) ,以解决人体内应用鼠单抗时的免疫原性问题 ,为进行抗 HCV的基因治疗研究开辟新途径。方法采用噬菌体表面展示技术 ,以重组的 HCV非结构蛋白 NS3为固相抗原 ,从噬菌体单链可变区抗体库中经过 5轮“吸附 -洗脱 -扩增”筛选过程 ,获得抗原结合活性较强的 HCVNS3人单链可变区抗体的阳性克隆 ,并对其进行免疫检测及序列测定。结果筛选出来的 Sc Fv片段具有抗 NS3的特异性。证实利用噬菌体抗体库技术 ,可以成功地获得 HCV NS3人单链抗体 Sc Fv的编码基因。结论筛选获得了 HCV非结构蛋白 NS3的特异性单链抗体的编码基因。Objective To screen and identify humanized single chain variable region antibody for hepatitis C virus non structural 3 protein. Methods Phage display technique was employed in the screening and identification of humanized single chain variable region(ScFv) antibody by using recombinant hepatitis C virus (HCV) non structural 3 protein as coating antigen. Results After 5 rounds of biopanning of a ScFv antibody phage library, 66 phage clones were evaluated by enzyme linked immunosorbent assay(ELISA).2 phage clones were selected from 66 clones according to the highest OD value in the ELISA identification and lowest cross reaction to the bovine serum albumin(BSA).The coding fragments of 2 ScFv were sequenced. After 5 rounds of biopanning of the ScFv phage library with initial 2.0×10 13 clones, the HCV NS3 antigen binding phage particles have been concentrated by 300 times. 66 phage clones were selected randomly for the subsequent ELISA screening and identification by using HCV NS3 and BSA as coating antigens, respectively. 2 phage clones were selected for their high A value in the ELISA identification and low cross reaction to BSA. The 2 inserts in the phage vector were sequenced and confirmed its ScFv characterization. Conclusion We have successfully screened and identified 2 clones of HCV NS3 specific ScFv. [

关 键 词：丙型肝炎病毒 噬菌体抗体 SCFV 非结构蛋白NS3 

分 类 号：R392-33[医药卫生—免疫学]