重组恶性症原虫红细胞结合抗原175功能区的制备及联合免疫.pdfVIP

• 14 • 第二军医大学学报 2004 Jan;2S (1) Acad J Sec Mil Med Univ ·伦著· 重组恶性症原虫红细胞结合抗原 175 功能区的制备及联合免疫 张冬梅，潘卫庆气第二军医大学基础医学部病原生物学教研室，上海 200433) [摘要] 町的:全合成恶性症原虫 eba-175Hf2 基因并在毕氏酵母中进行高效分泌表达，用该重组蛋白与我室研制的芜疾疫 苗侯选抗原 PfCP-2.9 进行联合免疫以观察是否存在竞争性免疫抑制。方自奋:采用不对称 PCR 法拼接合成 eba-175 [ f2 基因 (959 bp) ，用电转化方法将合成基因导入毕氏酵母中并进行诱导表达，采用离子交换层析和分子筛层析纯化 EBA-175HF2 蛋 白。结果:全合成 eba-175[f2 基因在毕氏酵母中高效分泌表达。重组 EBA一 175 nF2 和 PfCP-2.9 联合免疫后，以 ELISA 和 IFA 检测，针对 2 个蛋白联合免疫的抗体滴度明显高于 2 个蛋白单独免疫的滴度。结抢:重组 EBA-175HF2 和 PfCP-2.9 联 合免疫时不存在竞争性免疫抑制，这 2 个重要的芜疾疫苗候选抗原具有潜在联合应用前景。 [关键词〕 恶性芜原虫 ;175000 红细胞结合抗原;免疫性 [中图分类号] R 392.11 [文献标识码] A [文章编号] 0258-879X(2004)01-0014-04 Preparing domain n F2 fragment of Plasmodium falciparum 175 000 erythrocyte binding antigen and its combined immunization ZHANG Dong-M凹， P AN Wei-Qing (Department of Etiological Biology , College of Basic Medical Sciences , Second Military Medical U niversity , Shanghai 200433 , China) [ABSTRACT] Objective: To synthesize the eba-175 Hf2 gene of plωnodium falci归门im and express it in Pichia μstoris in the secreting form. The recombinant protein and PfCP-2. 9 protein were used for combined immunization to see if there is antigen competition. Methods: Asymmetric PCR-based method was utilized to synthesize the 959bp eba-175 Hf2 gene. Plas mid containing the synthetic gene was introduced into Pichia 户astoris by electroporation for inducible expression. The recom binant EBA-175 [ F2 protein was purified by ion-exchange and gel filtration chromatography. Results: The eba-175 Hf2 gene was successfully expressed in Pichia 户astoris in the secreting form. The antibody titers in mice immunized with the combined EBA-175 HF2 and Pf