肺癌细胞特异性结合肽的体内筛选及初步鉴定  

Selection and identification of the peptides specifically binding to lung cancer cell

作　　者：潘琴[1] 石磊[1] 潘雪刁[1] 巫玮[1] 金桂芳[1] 臧林泉[1] 

机构地区：[1]广东药学院,广东广州510006

出　　处：《山东医药》2008年第33期27-28,共2页Shandong Medical Journal

基　　金：国家自然科学基金资助项目(30572177);教育部重点课题基金资助项目(208105)

摘　　要：目的筛选能与肺癌细胞特异性结合的多肽并初步鉴定其亲和力和特异性。方法用人肺癌细胞A549使裸鼠致瘤,尾静脉注射肽库。取荷瘤鼠的肿瘤组织,用体内噬菌体展示技术进行筛选;经过3轮体内筛选,获得在肿瘤组织中富集的噬菌体;利用ELISA和细胞免疫化学DAB染色法初步鉴定各克隆亲和力、特异性;对阳性克隆行DNA测序,利用生物信息学对此多肽进行分析、比对。结果随机挑选的10个单克隆中,8个均对A549细胞具有较高亲和力,其中3号克隆亲和力最高,其可特异性结合肺癌细胞A549和NCI-H1299,且对A549的亲和力强于NCI-H1299。DNA测序结果显示该序列与美国国立生物技术信息中心(NCBI)GenBank DNA序列数据库和Swiss-Prot蛋白数据库中的已知基因和蛋白无同源性。结论筛选出了能与肺癌细胞特异性结合的多肽。Objective To obtain polypeptide which could specifically bind to lung cancer A549 cell line from peptide libraries, and indentify its affinity and specificity. Methods Nude mice were inoculated with lung cancer cells A549, then the Ph. D.-12 phage display peptide library was injected intravenously tail vein into mice. After 20 min the mice were sacrificed and the phage rescued from tumor tissues. The phage recovered from the tissues were amplified and purified then re-injected for next round screening.After screening for 3 rounds in vivo,the phage peptides that homed to the tumor tissues or cells were obtained. Then the positive and specific binding clones were identified by cell enzyme-linked immunosorbent assay （ELISA） and diaminobenzidine （DAB）, and these DNA sequences of identified clones was sequenced. Thus the amino acid sequence was deduced and analysed with bioin-formatics. Results After 3 rounds of panning, 10 phage clones were identified, 8 of them specially binds to the A549, then the 8 phage clones were identified by DAB,one of them specially binds to the A549 and NCI-H1299.Tbe amino acid sequence was analyzed in NCBI and Swiss-Prot, the results show that it had no similarity with the known proteins sequence in the database. Conclusion A peptide which is specific binding to lung cancer cell line A549 has been selected by phage display in vivo from peptide libraries.

关 键 词：噬菌体体内展示技术 噬菌体库 肺癌细胞 特异性结合肽 

分 类 号：Q7[生物学—分子生物学] R73[医药卫生—肿瘤]