摘要
目的 从噬菌体随机肽库中筛选出模拟弓形虫特异性抗原表位的短肽分子 ,并探讨其对弓形虫的保护性效果。方法 以纯化的弓形虫免疫兔血清IgG为配基 ,亲和筛选法富集特异性噬菌体 ,随机挑取噬菌体克隆用夹心ELISA法和Dot-ELISA法检测其特异性 ,混合 4个阳性克隆免疫小鼠 ,1月后攻击感染 ,观察小鼠发病时间和死亡时间。结果 经 3轮筛选 ,特异性噬菌体得到了有效富集 ,第 3轮洗脱噬菌体的产量为第 1轮的 16 7倍 ,随机挑取 2 7个噬菌体经Dot -Bloting和夹心ELISA法检测 ,有 2 4个能与弓形虫免疫兔血清及单克隆抗体特异反应。用致死量弓形虫攻击感染经阳性克隆免疫的小鼠 ,存活时间明显长于对照组。结论 免疫筛选噬菌体随机多肽库可获得具有保护性的弓形虫抗原表位 ,为弓形虫疫苗研制提供了新途径。
Aim To obtain the short peptides which mimic the antigen of Toxoplasma gondii through screening phage random peptide library with immunol rabbit sera.Methods Specific IgG was purified from immnol rabbit sera and used to immunoscreen a phage random peptide library of 12 amino acid residues displayed as a fusion to protein Ⅲ of filamentous phage M13 The positive clones were obtained by three rounds of biopanning,the reactivity of each clone binding to specific IgG was examined by Dot Bloting and Sandwich ELISA Four clones were selected and the mixture was prepared to immunize KunMing mice for three times and challenged with T gondii a month later,then observed the date of death Result The eluated phages could be enriched to 167 times through three rounds of biopanning 27phage clones from the third round of biopanning were randomly selected and 24clones of them could bind to the specific antibody of T gondii The survival time of the immunized mice with positive clones were much longer than the N S control group Conclusion The results indicate that we can obtain T gondii specific epitopes by immunoscreening phage random peptide library with immunol rabbit sera and the epitopes also have some protectivity to T gondii These findings provide the potential value for the preparation of the vaccine of T gondii
出处
《中国人兽共患病杂志》
CSCD
北大核心
2003年第4期42-44,共3页
Chinese Journal of Zoonoses
基金
湖南省"十五"重点科研资助项目
关键词
随机肽库
筛选
弓形虫
特异性抗原
表位
诱导
保护性免疫
Toxoplasma gondii
immunoscreening phage random peptide library
sandwich ELISA
protective immunonity
