人源性bFGF单链抗体的筛选、表达及鉴定  

Screening, expression and characterization of single chain Fv （scFv） against basic fibroblast growth factor

作　　者：王宏[1] 陶俊[2] 邓宁[1] 周丽君[3] 向军俭[1] 

机构地区：[1]暨南大学生命科学技术学院抗体工程中心,广州510632 [2]南方医科大学基础医学院免疫教研室 [3]解放军海军总医院中心实验科

出　　处：《中华微生物学和免疫学杂志》2010年第7期674-677,共4页Chinese Journal of Microbiology and Immunology

基　　金：基金项目：国家科技计划“863”专题课题（2009AA02Z112）

摘　　要：目的 用重组人碱性成纤维细胞生长因子（rhbFGF）筛选大容量人源噬菌体抗体库,获得能与bFGF特异性结合的人源性单链抗体（scFv）.方法 用rhbFGF对大容量人源噬菌体抗体库进行4轮＂吸附-洗脱-扩增＂筛选,并通过phage ELISA法对随机挑选的克隆进行抗原结合活性测定,得到的阳性克隆进一步进行原核可溶性表达及鉴定.结果 从104个随机挑选的克隆中,筛得39株抗bFGF的抗体,通过对抗体可变区基因进行多样性分析,发现存在8种不同抗体可变区基因,挑选其中8株phage ELISA显色最高的克隆进行酶切和测序鉴定,发现其中7株为正确的抗体基因序列,通过大肠杆菌成功的进行了scFv的可溶性表达,通过竞争ELISA发现其中一株scFv能够抑制bFGF与其高亲和力受体FGFR1βⅢC的结合.结论 通过筛选噬菌体抗体库获得到7株抗bFGF特异性抗体,其中一株抗体能够抑制bFGF与其高亲和力受体FGFR1的结合.Objective To obtain the specific human scFv basic fibroblast growth factor（bFGF）using phage antibody library technology. Methods The library was panned with human recombinant bFGF for 4 rounds. The antigen binding activities of random clones were tested by ELISA in order to select specific antibodies, which were then examined by DNA sequence analysis. Results The positive clone selected from the 104 random clones was able to bind bFGF specifically, while not able to bind other growth factors,such as aFGF, VEGF（vascular endothelial growth factor）. By competition ELISA assay we found one clone 44 could inhibit bFGF binding to FGFR1. Conclusion Seven specific human phage antibody against bFGF was obtained by phage display technique, one clone could inhibit bFGF binding to its high affinity receptor FGFR1.

关 键 词：噬菌体展示 抗体库 BFGF 人源性抗体 单链抗体 

分 类 号：R392[医药卫生—免疫学]