本文選題：HIV + 表位�。� 參考：《吉林大學(xué)》2009年博士論文

【摘要】： 為研制安全有效的艾滋病治療性疫苗,本研究在本室篩選出的一個(gè)較為成熟的多表位重組核酸疫苗(pVMEGNp24)的基礎(chǔ)上引入了4-1BBL及OX40L兩個(gè)協(xié)同刺激分子作為佐劑,構(gòu)建了pVMEGNp24-4-1BBL-OX40L以及pVMEGNp24-OX40L。在哺乳動(dòng)物細(xì)胞中表達(dá)的間接免疫熒光結(jié)果表明,所有構(gòu)建的重組質(zhì)粒編碼蛋白均得到了正確表達(dá)。 將己構(gòu)建好的DNA疫苗(pVMEGNp24-41BBL-OX40L),經(jīng)發(fā)酵罐建立發(fā)酵工藝,通過(guò)分子篩層析(Sepharose 6 FF)去除RNA,親和層析(PlasmidSelect Xtra)去除線性質(zhì)粒DNA,陰離子交換層析(SOURCE30Q)精純,以中空纖維交叉流系統(tǒng)濃縮至藥用劑量。經(jīng)過(guò)質(zhì)量鑒定結(jié)果表明,該純化質(zhì)粒疫苗臨床用質(zhì)粒DNA的質(zhì)量標(biāo)準(zhǔn),所建立的工藝可行。 將純化后的重組DNA(rDNA)疫苗質(zhì)粒通過(guò)導(dǎo)入人外周血來(lái)源的DC。并將轉(zhuǎn)染外源基因的成熟DC細(xì)胞與自體淋巴細(xì)胞共培養(yǎng)。免疫學(xué)指標(biāo)檢測(cè)結(jié)果表明:協(xié)同刺激分子在誘導(dǎo)細(xì)胞免疫應(yīng)答上作用強(qiáng)大,并且協(xié)同刺激分子的聯(lián)合應(yīng)用具有明顯的協(xié)同累積作用。 采用rDNA/rFPV(重組雞痘病毒)聯(lián)合免疫策略,免疫中國(guó)恒河猴,以人-猴艾滋病重組病毒SHIV-KB9攻毒后,分析疫苗免疫原性和攻毒保護(hù)效果。結(jié)果表明:協(xié)同刺激分子可以誘導(dǎo)免疫動(dòng)物產(chǎn)生更高水平的細(xì)胞及體液免疫反應(yīng)和較強(qiáng)的記憶性免疫反應(yīng)。SHIV-KB9病毒攻擊后,單協(xié)同刺激分子免疫組的動(dòng)物獲得的保護(hù)效果更好,單協(xié)同刺激分子疫苗是一個(gè)有潛力的候選疫苗。為HIV重組疫苗的研制與臨床前實(shí)驗(yàn)研究奠定了基礎(chǔ)。
[Abstract]:In order to develop a safe and effective AIDS therapeutic vaccine, a mature multiepitope recombinant nucleic acid vaccine (pVMEGNp24) was selected in this study. Two co-stimulant molecules, 4-1BBL and OX40L, were introduced as adjuvants to construct pVMEGNp24-4-1BBL-OX40L and pVMEGNp24-OX40L. The results of indirect immunofluorescence in mammalian cells showed that all the constructed recombinant plasmid encoded proteins were correctly expressed. The constructed DNA vaccine (pVMEGNp24-41BBL-OX40L) was fermented in fermenter. The RNAs were removed by Sepharose 6FF, the linear plasmid DNA was removed by PlasmidSelect Xtra, the anion exchange chromatography (SOURCE30Q) was purified and concentrated to medical dosage by hollow fiber cross flow system. The results of quality identification showed that the quality standard of plasmid DNA for clinical use of the purified plasmid vaccine was feasible. The purified recombinant DNA (rDNA) vaccine plasmid was transfected into DC1 from human peripheral blood. Mature DC cells transfected with exogenous genes were co-cultured with autologous lymphocytes. The results of immunological indices showed that the co-stimulator had a strong effect on inducing cellular immune response, and the combined application of co-stimulator had obvious synergistic cumulative effect. RDNA / rFPV (recombinant fowlpox virus) was used to immunize Chinese rhesus monkeys. The immunogenicity and protective effect of rDNA rFPV (recombinant fowlpox virus) vaccine against human monkey AIDS recombinant virus SHIV-KB9 were analyzed. The results showed that the co-stimulator could induce higher level of cellular and humoral immune response and stronger memory immune response of immunized animals, and the protective effect of the single co-stimulatory molecular immunization group was better than that of the control group. Single costimulatory molecular vaccine is a potential candidate vaccine. It lays a foundation for the development of recombinant HIV vaccine and preclinical experimental research.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2009
【分類號(hào)】：R392

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前1條

1 杜壽文;HIV復(fù)合表位核酸和痘苗病毒載體疫苗構(gòu)建及免疫原性研究[D];吉林大學(xué);2012年

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本文編號(hào)：2105444