B群链球菌42kD蛋白提取及其免疫学效果的初步探讨

发布时间：2018-01-10 19:26

  本文关键词：B群链球菌42kD蛋白提取及其免疫学效果的初步探讨　出处：《北京生物制品研究所》2005年硕士论文　论文类型：学位论文

  更多相关文章： GBS 结合疫苗 蛋白 免疫 主动保护 被动保护 交叉保护

【摘要】：B群链球菌(Group B Streptococcus)简称GBS,是引起新生儿肺炎、败血症和脑膜炎的主要病原菌。其多糖疫苗通常只产生较低的免疫活性,且多糖抗原在人体内易受降解,易导致免疫耐受等特点使得多糖疫苗的广泛应用具有一定的局限性。GBS外源蛋白(如:破伤风类毒素)结合疫苗虽然克服了B群链球菌荚膜多糖(CPS)疫苗的许多不足,但仍未解决血清型限制性的问题,而且单一载体蛋白不能对CPS提供足够的载体效应,增加载体蛋白的量所带来的毒性问题以及制备过程的复杂性也不容忽视。故现在寻求一种以GBS菌体免疫原性蛋白为基础的疫苗来解决上述问题。 本实验以一步层析法从GBS Ⅲ型菌32325株中提取42KD菌体蛋白,回收率为8%,等电点为6.6,纯度为100%,并初步鉴定为糖蛋白。它既可表达在细菌表面,又可分泌到菌体之外。以42KD蛋白不同剂量和针次免疫NIH成年小鼠,间接ELISA法检测小鼠血清IgG抗体滴度。结果显示:不同剂量免疫均能诱导小鼠产生相应IgG抗体,其效价最高可达1:24576,说明42KD蛋白具有很好的免疫原性。选择最佳免疫条件免疫小鼠,然后腹腔攻击GBSⅢ型32325菌株,观察主动免疫和被动免疫保护效果。结果显示:试验组成年小鼠的免疫保护指数为100,乳鼠的免疫保护指数为40,表明试验组较阴性对照组耐受活菌攻击量分别大100倍和40倍。用1个LD_(50)和10个LD_(50)剂量菌液攻击试验组成年小鼠后存活率均可达100%,阴性对照组分别为50%和37.5%;在被动免疫保护性试验中,用1个乳鼠的LD_(50)剂量攻击,试验组乳鼠存活率可达100%,而阴性对照组为50%,试验组存活率显著高于阴性对照组。这些结果表明42KD蛋白对成年小鼠和乳鼠均诱导保护免疫。不同剂量免疫保护试验结果说明:在一定范围内,免疫剂量与血清IgG抗体应答水平呈正相关;小鼠血清IgG抗体滴度与保护力正相关。用肺炎链球菌Pn6B:CMCC 31492株和Pn19F:CMCC 31699株作交叉保护性试验,结果显示,42KD蛋白对肺炎链球菌不具有交叉保护作用。
[Abstract]:Group B streptococcus group B Streptococcus, referred to as GBS, is the cause of neonatal pneumonia. The main pathogen of septicemia and meningitis. The polysaccharide vaccine usually produces only low immune activity and the polysaccharide antigen is easily degraded in human body. It is easy to lead to immune tolerance and so on, which makes the wide application of polysaccharide vaccine have certain limitations. GBS exogenous protein (such as tetanus toxoid) conjugate vaccine has overcome group B streptococcus capsule polysaccharide (CPS). Many of the shortcomings of the vaccine. However, the problem of serotype restriction has not been solved, and the single carrier protein can not provide enough carrier effect to CPS. The toxicity caused by increasing the amount of carrier protein and the complexity of the preparation process can not be ignored. Therefore, a vaccine based on the immunogenicity protein of GBS cell is now sought to solve the above problems. In this experiment, 42KD bacterial protein was extracted from 32325 strains of GBS 鈪，

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