Serpin多抗制备和产Serpin的双歧杆菌菌株的初步筛选

Preparation of Polyclonal Antibodies of Serpin and Preliminary Screening of the Bifidobacteria Species of Yielding Serpin

DOI：

中文关键词: 长双歧杆菌WBL001 serpin 纯化 抗血清 斑点杂交

英文关键词: Bifidobacterium longum WBL001 serpin purification antiserum colony immunoblotting detection

基金项目:国家863计划项目(2008AA10Z337);国家自然科学基金项目(30900038,31000048);江西省教育厅科研基金(GJJ10379)

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中文摘要:

从长双歧杆菌WBL001基因组中克隆了serpin基因,构建重组Serpin蛋白的原核表达体系pBX2-serpin,纯化表达产物作为抗原,免疫小鼠制备Serpin的多克隆抗体。采用偶联Serpin的磁珠纯化多克隆抗体,并探究了不同溶剂对抗体洗脱效率的影响。纯化的多抗用于斑点杂交筛选产Serpin蛋白的菌株。结果表明:磁珠纯化多克隆抗体以1 mol/L NaOH的洗脱率最高,达50%;纯化后的Serpin抗体消除了与部分乳酸菌和致病菌的非特异性反应;采用斑点杂交方法从11株双歧杆菌中筛选到婴儿双歧杆菌WBAN07具有类Serpin蛋白。

英文摘要:

Serpin was amplified from the genome of Bifidobacterium longum WBL001.Recombinant plasmid pBX2-serpin was constructed and expressed in Escherichia coli for Serpin production.The recombinant protein was purified and used as antigen for mice immunization.Antiserum was purified specifically by Serpin-coupled magnetic beads.The antibody eluting was optimized by using different eluents.Serpin-like protein was screened out from 11 Bifidobacteria strains by colony immunoblotting.The result indicated that 1 mol/L NaOH is the best fluent for antibody elution from magnetic beads(about 50% recovery).Non-specific reactions of antiserum with lactic acid bacterium and pathogens were eliminated after purification.Serpin-like protein had been found in B.infantis WBAN07 from 11 Bifidobacteria strains by colony immunoblotting.

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