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Construction of multivalent recombinant Aβ42 DNA vaccine and induction of Aβ antibody
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- Author:
- No author available
- Journal Title:
- JOURNAL OF THE FOURTH MILITARY MEDICAL UNIVERSITY
- Issue:
- 10
- DOI:
- 10.3321/j.issn:1000-2790.2005.10.001
- Key Word:
- 淀粉样β蛋白;疫苗,DNA;Aβ抗体
Abstract： 目的: 构建β-淀粉样蛋白(Aβ42)的多价重组DNA疫苗,确定是否诱导较高滴度Aβ抗体的产生.方法: PCR法从Tg2576转基因鼠基因组DNA中扩增人Aβ42基因片段;在Aβ42基因的5′端引物延伸连接IgG к轻链信号肽序列;利用GSGGSG linker基因串连两段Aβ42基因片段;将产物克隆入pcDNA3.1表达载体,分别构建pcDNA3.1-Aβ42,一价分泌型pcDNA3.1-sAβ42,二价分泌型pcDNA3.1-s2×Aβ42载体;转染COS-7细胞,免疫印迹检测Aβ42的表达;im接种BALB/c鼠,ELISA法测定抗体效价;免疫组织化学和Aβ42肽抗原的中和实验检测抗体与Tg2576鼠脑切片中的老年斑的结合.结果: 测序证实所构建的pcDNA3.1-Aβ42,pcDNA3.1-sAβ42,pcDNA3.1-s2×Aβ42载体插入序列正确;Western Blotting证实在COS-7细胞中表达相应的蛋白.经6轮,22 wk加强免疫后,pcDNA3.1-Aβ42疫苗不能产生Aβ抗体、pcDNA3.1-sAβ42接种后产生较低的Aβ抗体(<1:1000),pcDNA3.1-s2×Aβ42第1次加强免疫后即产生抗体,抗体滴度随接种次数增多而升高,22 wk时抗体平均滴度达1:5000以上,可以与转基因鼠Tg2576脑切片中的老年斑结合,并能被Aβ42肽抗原特异性中和.结论: 二价分泌型pcDNA3.1-s2×Aβ42疫苗,可在动物体内产生高效价的特异Aβ抗体.
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