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目的 :利用纯化的抗大肠杆菌L2 6 30多抗筛选噬菌体环七肽库 ,以获得可模拟脂多糖 (LPS)表位的短肽克隆。方法 :以亲和层析纯化的抗大肠杆菌L2 6 30多克隆抗体为靶分子 ,筛选噬菌体随机环七肽库 ,用双夹心ELISA和竞争抑制ELISA鉴定阳性噬菌体克隆的抗原性。结果 :对噬菌体环肽库进行 3轮筛选后 ,随机挑选 2 0个克隆 ,经鉴定其中 12个可与抗L2 6 30抗体结合 ,即为阳性克隆 ;其中有 5个阳性噬菌体克隆表现出与抗鼠伤寒沙门氏菌LPS多抗结合的活性 ,提示这 5个噬菌体展示的短肽具有模拟大肠杆菌LPS及鼠伤寒沙门氏菌LPS共同表位的性质。经DNA序列分析显示 ,其中 8个克隆的氨基酸序列具有X DGLL XX或X EDGLL X保守序列 ,其余 4个克隆的序列均不相同。结论 :筛选获得的噬菌体环七肽克隆具有模拟大肠杆菌LPS表位的活性 ,为大肠杆菌L2 6 30多表位模拟短肽。其中 5个阳性噬菌体克隆短肽具有模拟大肠杆菌LPS及鼠伤寒沙门氏菌LPS共同表位的活性
OBJECTIVE: To screen a phage cloning library using purified anti-Escherichia coli L2 6 30 polyclonal antibody to obtain short peptide clones that mimic the lipopolysaccharide (LPS) epitopes. Methods: The anti-E.coli L2 6 30 polyclonal antibody purified by affinity chromatography was used as a target. The phage randomized cyclic heptapeptide library was screened. The antigenicity of positive phage clones was identified by double-sandwich ELISA and competitive inhibition ELISA. RESULTS: After three rounds of screening of the phage cyclic peptide library, 20 clones were randomly selected and 12 of them were identified as positive clones by binding to the anti-L2 6 30 antibody. Among them, 5 positive phage clones showed resistance to anti- Salmonella typhimurium LPS polyclonal anti-binding activity, suggesting that the five phage displayed short peptides with the simulation of E. coli LPS and Salmonella typhimurium LPS common epitope properties. DNA sequence analysis showed that the amino acid sequence of the eight clones had the conserved sequence of X DGLL XX or X EDGLL X and the sequence of the other four clones was different. CONCLUSION: The cloned phage cloned heptapeptide clone has the activity of simulating the LPS epitope of E. coli and is an E. coli L2 6 30 multi-epitope mimic peptide. Among them, five positive phage clone short peptides had the activity of mimicking E. coli LPS and S. typhimurium LPS common epitope