全人源抗鼻咽癌噬菌体单链抗体的筛选与鉴定  

Screening and characterization of human scFv antibodies against nasopharyngeal carcinoma

作　　者：李艳东[1] 谢平丽[1] 王甲甲[1] 李跃辉[1] 胡锦跃[1] 李官成[1] 

机构地区：[1]中南大学湘雅医学院肿瘤研究所,湖南长沙410078

出　　处：《中国医师杂志》2009年第5期577-580,共4页Journal of Chinese Physician

基　　金：“863”项目（2007AA021809,2007AA021811）

摘　　要：目的从全人源抗鼻咽癌噬菌体抗体库中筛选特异性单链抗体（ScFv），并对其特异性进行鉴定。方法通过噬菌体表面展示技术把ScFv表达在噬菌体表面，以鼻咽癌细胞作为抗原，用抗原递减法，通过“吸附-洗脱-扩增”过程筛选并富集特异性抗体，及ELISA筛选，获得特异阳性克隆进行免疫组化鉴定并测序。结果通过对抗体库进行三轮正负淘洗和富集后，随机挑选4212个克隆进行ELISA，发现3个克隆对CNE2呈强阳性反应，而与人正常细胞系HUVEC等呈弱阳性反应或不反应。对克隆HNSA033进一步进行免疫细胞化学验证，结果与ELISA反应一致；免疫组织化学鉴定表明克隆HNSA033与鼻咽癌组织和鼻咽组织阳性率的差别有统计学意义。结论通过淘选富集、ELISA和免疫化学鉴定获得特异性较强的噬菌体克隆，为鼻咽癌发病机制的研究和临床诊断以及治疗奠定了基础。Objective To screen the anti-nasopharyngeal carcinoma scFv from a human anti-nasopharyngeal carcinoma single-chain phage antibody library, and identify its characteristies. Methods The single-chain phage antibody library was subjected to three rounds of positive and negative cell panning and enrichment, and then it was selected by ELISA. The binding specificity of phage antibodies with nasopharyngeal carcinoma cells was confirmed by immunohistochemistry. Results After panning, enrichment and testing by ELISA, 3 phage antibody clones reacting with CNE2 more strongly than HUVEC and NP69 were picked out from 4212 clones. One clone, HNSA033, was further analyzed after DNA sequencing. The results of immunohistoehemistry with cultured cells were similar to those of ELISA. HNSAO33 specifically reacted to nasopharyngeal carcinoma cells in most human nasopharyngeal carcinoma tissue seetians except a few human normal nasopharyngeal tissue sections. The distinction of positive rates was of a great statistical signifieance. Conclusion ELISA and immunohistochemistry results confirmed HNSA033 specifically bind with nasopharyngeal carcinoma cells. The seFv fragment against nasopharyngeal eareinoma may be further developed and applied in clinical diagnosis and therapy of nasopharyngeal carcinoma.

关 键 词：细菌噬菌体/免疫学 抗体 单克隆/生物合成 鼻咽肿瘤/免疫学 

分 类 号：R739.63[医药卫生—肿瘤]