Li Xun, Xue Cai-fang, Liu Zhong-xiang, Wang Xian-feng, Ding Jin, Lei Jun-chuan, Zhen Rong-fen
Department of Pathobiology, Fourth Military Medical University, Xi'an 710032.
Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2003;21(2):80-3.
To determine the role of putative apical membrane antigen (AMA)1 domains in inducing protective immunity and to provide basis for selection of vaccine applicable segments.
Encoding gene segments of AMA1 were amplified and cloned into pET prokaryotic expression vectors. Recombinant proteins were expressed and purified. Groups of BALB/c mice were immunized by using recombinant protein in Freund's adjuvant, and the IgG titer and specificity of the immune sera were analyzed by IFA and Western blotting. Efficiency of the immune sera in inhibiting Plasmodium falciparum in vitro growth was evaluated.
Recombinant AMA1 fragments including the entire ectodomain E and subdomain I + II, I, II and III were successfully expressed and purified. Different levels of antibody were induced in mice by individual proteins and all the immune sera recognized native antigen in the parasites. Sera from protein E and I + II immunized mice inhibited the growth of parasites.
The integrality of the ectodomain of AMA1 determines the conformation of the protective antibody epitopes, and these protective epitopes distribute mainly in the subdomain I.
确定假定的顶端膜抗原(AMA)1结构域在诱导保护性免疫中的作用,并为选择适用的疫苗片段提供依据。
扩增AMA1的编码基因片段并克隆到pET原核表达载体中。表达并纯化重组蛋白。用弗氏佐剂中的重组蛋白免疫BALB/c小鼠组,通过间接荧光抗体试验(IFA)和蛋白质印迹法分析免疫血清的IgG滴度和特异性。评估免疫血清在体外抑制恶性疟原虫生长的效率。
成功表达并纯化了包含整个胞外结构域E和亚结构域I+II、I、II和III的重组AMA1片段。单个蛋白在小鼠中诱导了不同水平的抗体,并且所有免疫血清都识别寄生虫中的天然抗原。来自用蛋白E和I+II免疫的小鼠的血清抑制了寄生虫的生长。
AMA1胞外结构域的完整性决定了保护性抗体表位的构象,并且这些保护性表位主要分布在亚结构域I中。