【摘要】： 目的構(gòu)建日本血吸蟲多價膜錨定表達疫苗pIRES-Sj97-Sj14-Sj26,并研究其免疫原性。 方法采用分子克隆技術(shù),構(gòu)建日本血吸蟲脂肪酸結(jié)合蛋白(Sj14)、GST(Sj26)和副肌球蛋白(Sj97)的跨膜共表達質(zhì)粒pIRES-Sj97-Sj14-Sj26,轉(zhuǎn)染Hela細胞,通過RT-PCR法檢測Sj14、Sj26和Sj97mRNA的表達,間接免疫熒光檢測Sj26蛋白的表達。用此疫苗免疫BALB/c小鼠,同時設(shè)置生理鹽水對照組、空白質(zhì)粒對照組、pIRES-Sj26組和pIRES-Sj14-Sj26組。用ELISA法檢測免疫小鼠血清中的總IgG抗體和小鼠脾細胞分泌IFN-γ的水平。以MTT法檢測淋巴細胞的增殖情況。并以FCM分析脾細胞亞群。 結(jié)果經(jīng)瞬時轉(zhuǎn)染Hela細胞證明質(zhì)粒pIRES-Sj97-Sj14-Sj26能在體外進行表達。免疫小鼠后ELISA法檢測抗體結(jié)果表明pIRES-Sj97-Sj14-Sj26能夠較各對照組誘導產(chǎn)生高水平的IgG抗體(P0.01,P0.05).該組的脾淋巴細胞刺激指數(shù)較各對照組也有顯著增高(P0.01),IFN-γ的水平與空白對照組和空載體組有顯著差異(P0.01). CD4+和CD8+T細胞含量百分比有明顯增高(P0.05)。 結(jié)論pIRES-Sj97-Sj14-Sj26疫苗具有較強的免疫原性,能明顯增強BALB/c小鼠的免疫應答反應.為該疫苗抗感染效應的研究奠定了基礎(chǔ),并為日本血吸蟲病的防治尋求一種新的多基因組合疫苗的免疫策略。
[Abstract]:Objective to construct and study the immunogenicity of the multivalent membrane anchored pIRES-Sj97-Sj14-Sj26, vaccine of Schistosoma japonicum. Methods the transmembrane coexpression plasmid pIRES-Sj97-Sj14-Sj26, of fatty acid binding protein (Sj26) and accessory myosin (Sj97) of Schistosoma japonicum was constructed by molecular cloning technique. The expression of Sj14,Sj26 and Sj97mRNA was detected by RT-PCR method, and the expression of Sj26 protein was detected by indirect immunofluorescence. BALB/c mice were immunized with this vaccine, and the control group of normal saline, the control group of blank plasmid pIRES-Sj26 and the group of pIRES-Sj14-Sj26 were set up at the same time. The total IgG antibody in serum of immunized mice and the level of IFN- 緯 secreted by spleen cells of mice were detected by ELISA method. The proliferation of lymphocytes was detected by MTT method. Spleen cell subsets were analyzed by FCM. Results transient transfection of Hela showed that plasmid pIRES-Sj97-Sj14-Sj26 could be expressed in vitro. After immunizing mice, the results of ELISA assay showed that pIRES-Sj97-Sj14-Sj26 could produce higher level of IgG antibody than that of control group (P0.01P 0.05). The level of IFN- 緯 was significantly higher in this group than that in the control group (P0.01), and there was a significant difference between the control group and the blank carrier group (P0.01). The percentage of CD4 and CD8 T cells was significantly increased (P0.05). Conclusion pIRES-Sj97-Sj14-Sj26 vaccine has strong immunogenicity and can obviously enhance the immune response of BALB/c mice. It lays a foundation for the study of the anti-infective effect of the vaccine and seeks a new immunization strategy for the prevention and control of schistosomiasis japonicum.
【學位授予單位】：華中科技大學
【學位級別】：碩士
【學位授予年份】：2007
【分類號】：R392

【參考文獻】

相關(guān)期刊論文 前10條

1 郝文波,羅樹紅;核酸疫苗——寄生蟲疫苗的新希望[J];國外醫(yī)學(寄生蟲病分冊);1996年05期

2 李萍,嚴家新;DNA疫苗研究進展[J];國外醫(yī)學.預防.診斷.治療用生物制品分冊;1995年03期

3 張冉;血吸蟲病DNA疫苗的研究進展[J];中國熱帶醫(yī)學;2004年01期

4 李柳哲,石佑恩,江侃,寧長修,鄧偉文;日本血吸蟲多價DNA疫苗pBK-Sj26 (Sj32)-Sj23免疫效果的觀察[J];同濟醫(yī)科大學學報;2001年05期

5 李柳哲,石佑恩,寧長修,鄧偉文,韓家俊;日本血吸蟲混合DNA疫苗免疫效果觀察[J];中國血吸蟲病防治雜志;2001年04期

6 馮新港,余新炳,吳忠道,周俊梅;日本血吸蟲SjFABPc重組質(zhì)粒裸DNA免疫小鼠的研究[J];中國血吸蟲病防治雜志;2001年05期

7 朱蔭昌,任建功,司進,D.A.Harn,余傳信,梁幼生,殷旭仁,徐明,許永良;日本血吸蟲SjCTPI和SjC23DNA疫苗聯(lián)合免疫保護作用的研究[J];中國血吸蟲病防治雜志;2002年02期

8 余傳信,朱蔭昌,殷旭仁,任建功,司進,許永良,沈林南,何偉;日本血吸蟲脂肪酸結(jié)合蛋白(SjC FABP)核酸疫苗誘導小鼠免疫保護作用研究[J];中國血吸蟲病防治雜志;2002年03期

9 鄭江;中國血吸蟲病防治現(xiàn)狀及展望[J];中國血吸蟲病防治雜志;2003年01期

10 司進,朱蔭昌,Harn DA,殷旭仁,余傳信,何偉,任建功,華萬全;日本血吸蟲中國大陸株TPI DNA疫苗誘導BALB/c小鼠保護性免疫力的研究[J];中國寄生蟲病防治雜志;2001年03期

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