应用噬菌体随机肽库研究细粒棘球蚴抗原模拟表位

Screening of Mimic Antigen Epitopes of Echinococcus granulosus by Phage-displayed Peptide Library

用抗细粒棘球蚴B抗原(EgB)多克隆抗体筛选噬菌体随机7肽库。经过5轮筛选后,噬菌体回收率从第1轮的4.15×10-5增加到第5轮的4.30×10-2,说明阳性克隆得到富集。随机挑取60个蓝色噬菌斑进行扩增,对其核苷酸序列进行测定分析并与EgB进行同源性比较。采用ELISA法对阳性噬菌体与抗EgB多克隆抗体结合特性进行检测,共检测出45个与抗EgB多克隆抗体结合的克隆株(阳性率为75%),其递呈的七肽序列与EgB的同源性低。采用ELISA法检测所选多肽对棘球蚴病所致过敏性休克患者抗血清的反应性,结果显示,细粒棘球蚴病患者抗血清与45个阳性噬菌体克隆反应的吸光度(A410值)比其与7肽库反应的A410值大2倍以上,该45个阳性噬菌体克隆与EgB多克隆抗体的结合是特异的。提示成功分析细粒棘球蚴囊液B抗原肽表位以及模拟表位。

The 7-mer phage random peptide libraries was screened with the specific anti-EgB polyclonal antibody. Sixty clones picked randomly were deduced by DNA sequencing and their amino acid sequences were compared with EgB. After 5 rounds of panning, the phage recovery rate increased from 4.15×10-5 to 4.30×10-2. 45 out of 60 selected clones were ELISA positive by anti-EgB polyclonal antibody with a positive rate of 75%, 45 positive clones were sequenced and the predicted amino acid sequences showed low homology to EgB. The reactivity of selected peptide and sera from patients of anaphylactic shock induced by Echinococcus granulosus was detected by ELISA. The result showed that the absorbance value （A410） of 45 postive clones detected by using serum samples from patients was twice higher than that by using 7 peptide library. The 45 positive clones exhibited specific binding to anti-EgB antibody. The peptides screening from 7-mer phage random peptide libraries may be the mimic epitopes of anaphylactic shock induced by E. granulosus.