鲍曼不动杆菌烈性噬菌体的分离与纯化

利用柱层析方法,纯化鲍曼不动杆菌（Acinetobacter baumannii）烈性噬菌体AB1。首先采用聚乙二醇6000沉淀方法,初步分离裂解液中的噬菌体,噬菌体纯度由6.1×1010 pfu/mg提高到37×1010 pfu/mg,噬菌体回收率为58.8%,蛋白质去除率为90.6%;噬菌体粗提样品经Sepharose 4B凝胶过滤层析柱进一步纯化,纯度提高到73×1010 pfu/mg,噬菌体回收率为95.7%,蛋白质去除率为48.1%;收集的噬菌体样品最后经DEAE-52阴离子交换层析柱处理,噬菌体纯度为40×1010 pfu/mg,回收率为50.8%,蛋白去除率15.6%。内毒素分析结果显示,Sepharose 4B凝胶过滤层析纯化的噬菌体样品中,内毒素含量为443.8 EU/mg,而DEAE-52阴离子交换层析纯化的噬菌体样品中,内毒素含量为544.4 EU/mg。实验结果显示,PEG沉淀方法与Sepharose 4B凝胶过滤方法能够有效地提高噬菌体纯度,而DEAE-52阴离子交换层析则不能提高噬菌体的纯度,也无法有效地去除样品中的内毒素。

Purification of Acinetobacter baumannii bacteriophage

The virulent bacteriophage AB1 specific to an Acinetobacter baumannii clinical strain was purified with combined chromatography methods.Polyethylene glycerol 6000（PEG6000） was used to precipitate phage particles from lysate.Analysis of the phage suspension showed that the purity of phage was increased from 6.1 × 1010 to 37 × 1010 pfu/mg,phage recovery rate was 58.8%,and protein removal rate was 90.6%.Phage sample was subsequently purified by Sepharose 4B gel filtration chromatography.Analysis of the phage samples showed that the purity of phage was increased to 73 × 1010 pfu/mg,phage recovery rate was 95.7%,and protein removal rate was 48.1%.DEAE-52 anion-exchange chromatography was adopted to purify the phage samples,analysis showed that the phage purity was decreased to 40 × 1010 pfu/mg,phage recovery rate was 50.8%,and protein removal rate was 15.6%.Endotoxin contents of the purified phage samples were also investigated,443.8 EU/mg in Sepharose 4B gel filtration treated sample and 544.4 EU/mg in DEAE-52 anion-exchange treated sample.Experimental results indicated that PEG precipitation method and Sepharose 4B gel filtration method could efficiently increased the phage purity,while DEAE-52 anion-exchange chromatography was not suitable in purifying phage particles.