抗大肠杆菌LPS多克隆抗体的制备与LPS模拟肽的筛选

目的获得抗大肠杆菌脂多糖(LPS)多克隆抗体与模拟LPS表位的线性噬菌体展示肽克隆。方法制备兔抗鼠大肠杆菌抗血清,鉴定抗血清效价。以亲和纯化的多克隆抗体为靶,筛选噬菌体随机线性十二肽库。双夹心ELISA和竞争抑制ELISA鉴定阳性噬菌体克隆。结果兔抗血清能与大肠杆菌LPS反应。用亲和层析纯化的多克隆抗体为靶分子进行三轮筛选,随机挑选17个克隆,其中6个克隆显示与多抗结合。大肠杆菌LPS可抑制阳性噬菌体克隆与多抗结合,所有克隆的IC50(达到50%抑制率的LPS浓度)为250ng/ml。结论获得能与大肠杆菌LPS反应兔多克隆抗体,筛选得到可模拟大肠杆菌LPS表位的噬菌体展示线性十二肽。

Preparation of Polyclonal Anti-E. coli LPS Antibodies and Screening of Mimotopes of LPS from Liner 12-mer Phage Displayed Peptide Library

Objective To prepare the polyclonal anti-E. coli LPS antibodies and screen mimotopes of LPS from liner 12-mer phage displayed peptide library. Methods Rabbits were immunized intradermally with E. coli 2755. The antisera were tested for reaction to LPS of E. coli 2755 by ELISA. Mimotopes of LPS were screened from liner 12-mer phage displayed peptide library by biopaning using purified antibodies against LPS of E. coli 2755, and the antigenicity of selected clones was identified by double-sandwich ELISA and competitive ELISA. Results Rabbit antisera could react with E. coli LPS. After the third-round screening, 6 of 17 randomly selected phage clones were identified as positive that can bind to polyclonal antibodies. The IC50 of LPS against the binding between the positive phage clones and anti-LPS antibodies was 250ng/ml. Conclusion Mimotope clones of LPS screened from phage displayed peptide library can bind to anti-LPS antibodies, suggesting the peptide displayed on phage can mimic epitope of E. coli LPS.