用抗HCV多抗从随机12肽库中筛选抗原表位  

Biopanning of HCV antigen epitopes from a random 12-peptide library by anti-hepatitis C virus polyclonal antibody

作　　者：王敏[1] 李先平[1] 王庆林[2] 汤兰桂[2] 

机构地区：[1]中南大学湘雅二医院检验科,长沙410011 [2]湖南师范大学医学院医学检验系,长沙410013

出　　处：《中南大学学报（医学版）》2010年第3期236-240,共5页Journal of Central South University ：Medical Science

摘　　要：目的:用丙型肝炎患者血清中抗丙型肝炎病毒(HCV)抗体从噬菌体随机12肽库中筛选HCV抗原表位。方法:将丙型肝炎患者血清混合,提取纯化的IgG作为固相配基筛选噬菌体随机12肽库,按吸附-洗脱-扩增的淘洗过程进行3轮筛选,随机挑取噬菌体克隆用ELISA法检测其特异性,对4个克隆进行测序。并用ELISA法检测噬菌体克隆的诊断价值。结果:3轮筛选的投入产出比逐轮升高,回收率从(4.6×10-4)%增加到(5.3×10-2)%,具有良好的富集效果。对从第3轮洗脱物中挑选出的18个克隆进行结合试验,发现所挑的克隆与丙型肝炎患者的多克隆抗体有较强的结合力。其中4个克隆测序显示为同一克隆(命名为C1),其外源插入肽为GSMSPYVRWYTP。用C1检测20例丙型肝炎患者血清的检出率为85.0%。结论:成功地用噬菌体12肽库对丙型肝炎患者血清进行了模拟肽筛选,且得到的模拟肽分子C1具有一定的诊断价值。Objective To analyze the epitopes of anti-hepatitis C virus（HCV）antibodies by peptide library biopanning.Methods Phage random peptide library of 12 amino acids was immunoscreened with purified IgG from the sera of hepatitis C patients.Positive clones which were obtained after 3 rounds of biopanning were detected by ELISA and 4 of them were sequenced.Results After 3 rounds of screening,the radio of output to input increased from（4.6×10-4）% to（5.3×10-2）%,meaning the enrichment was effective.At the third round of screening,all the selected clones proved to specifically react with the sera for immunoscreening.Four positive phage clones were sequenced,which shared a very conservative sequence and was named as C1.Its inserting sequence in the coat protein III was deduced to be GSMSPYVRWYTP,and the positive rate of C1 reacted with 20 cases of HCV patients was 85%.Conclusion The antigen-mimic peptide C1 is successfully screened from 12 random phage peptide library and it has some diagnostic value.

关 键 词：噬菌体随机肽库 模拟表位 丙型肝炎病毒 

分 类 号：R392[医药卫生—免疫学]