摘要
本文发展了一种从百日咳杆菌Ⅰ相菌株中纯化６９ＫＤａ外膜蛋白的简易方法，将细菌体经加热浸提、乙醇沉淀蛋白、ＤＥＡＥ－ＳｅｐｈａｄｅｘＡ＿（５０）柱层析精制而成。用ＳＤＳ－ＰＡＧＥ、免疫印迹、光密度仪扫描分析，证明纯化制剂为均一的、特异性的６９ＫＤａ外膜蛋白，其收率为５４．２％，纯度达９９．２％，每微克６９ＫＤａ蛋白制。剂中内毒素含量低于０．８５ＥＵ；ＰＴ残留量小于０．１０５ｎｇ。ｆＬ６９ＫＤａ蛋白抗血清能凝集含有３因子凝集原的百日咳菌株。用纯化的６９ＫＤａ蛋白免疫小鼠，能够保护小鼠抵抗致死剂量的１８３２３菌株的脑内攻击，与ＰＴ、ＦＨＡ组成的无细胞百日咳菌苗的效力，较含ＰＴ和ＦＨＡ的组分菌苗为高。表明６９ＫＤａ蛋白与ＰＴ、ＦＨＡ显示协同保护作用，有望成为无细胞百日咳菌苗的另一候选组分。
A simplyfied method was developed to purify the 69KDa outer membrane protein from phase I Bordetella pertussis. Harvested bacterial cells were extracted by heating, precipitated with ethanol, followed by DEAE-Sephadex A_(50) chromatography. The purified preparation showed a specifical homogenous 69KDa outer membrane protein on SDS-PAGE, densitometric scanning and immunoblotting, its recovery was 54. 2%, the purity reached 99. 2%. The residue of endotoxin and PT were less than 0. 85EU and 0. 105ng per ug 69KDa protein respectively. Anti69KDa protein antiserum could agglutinate all B. pertussis cells containing serotype 3 agglutinogen. Active immunization of mice with 69KDa protein could protect mice from lethal intracerebral challenge with virulent strain. The relative potency of three components pertussis vaccine containing PT, FHA and 69KDa protein was higher than that of PT and FHA two components pertussis vaccine. It indicated 69KDa protein has a synergistic effect with PT and FHA. So 69KDa protein Could be considered as an additional component of candidate of acellular pertussis vaccine.
出处
《微生物学免疫学进展》
1995年第2期65-72,共8页
Progress In Microbiology and Immunology
关键词
百日咳杆菌
69KDa外膜蛋白
菌苗
分离
提纯
Bordetella pertussis Acellular pertussis vaccine 69KDa outer membrane protein
