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  本文選題：HIV-1　切入點：共轉(zhuǎn)染　出處：《吉林大學(xué)》2006年碩士論文

【摘要】：艾滋病的流行已成為人類健康和社會穩(wěn)定的大敵,我國已進入了艾滋病快速增長期,因此迫切需要一種有效的疫苗來阻止艾滋病的流行�，F(xiàn)在,已經(jīng)設(shè)計出很多能誘導(dǎo)廣泛有效中和抗體的候選疫苗。疫苗研究的進展要求在體外準確計量中和抗體的中和水平,以評價抗體和疫苗在實驗動物及人體內(nèi)的保護效果,所以,傳統(tǒng)的中和實驗技術(shù)必須進行改進,以滿足理論研究進展的需要。一個準確、可重復(fù)性的體外中和實驗對于研究病毒中和的機制,評價候選疫苗誘導(dǎo)中和反應(yīng)能力都是至關(guān)重要的。 本論文研究是利用熒光素酶作為報告基因,表達假病毒進行中和抗體評價,即構(gòu)建中國流行株各亞型HIV-1 envelope基因的真核表達質(zhì)粒和帶有熒光素酶報告基因的pNL-Luc-E—R—質(zhì)粒共轉(zhuǎn)染293T細胞上,培養(yǎng)收獲含有假病毒。假病毒和抗體或血清共孵育后感染表達CD4受體和一個輔助受體(CCR5 /CXCR4)的HOS細胞,培養(yǎng)3天后,用PBS洗滌細胞,將細胞裂解,檢測熒光素酶含量。并進一步建立穩(wěn)定表達HIV-1假病毒的細胞系,批次大量獲取假病毒,以適用于大批量的疫苗臨床受檢樣品。這種方法大大降低了傳統(tǒng)方法中宿主細胞( PBMC)的差異而引起的結(jié)果變異,是一種具有可重復(fù)性的極有發(fā)展前景的體外中和實驗方法。
[Abstract]:The AIDS epidemic has become a major enemy of human health and social stability. Our country has entered a period of rapid growth of AIDS. Therefore, an effective vaccine is urgently needed to stop the spread of AIDS. Many candidate vaccines have been designed to induce a wide range of effective neutralizing antibodies. Progress in vaccine research requires that neutralization levels of neutralizing antibodies be accurately measured in vitro to evaluate the protective effects of antibodies and vaccines in laboratory animals and humans, so, The traditional neutralization experiment must be improved to meet the needs of theoretical research. An accurate and repeatable in vitro neutralization experiment is needed to study the mechanism of virus neutralization. Evaluating the ability of candidate vaccines to induce neutralization is essential. In this study, luciferase was used as a reporter gene to evaluate the neutralization antibody of pseudovirus. The eukaryotic expression plasmid of HIV-1 envelope gene of Chinese epidemic strain and pNL-Luc-E-R- plasmid with luciferase reporter gene were cotransfected into 293T cells. Cultured and harvested HOS cells containing pseudovirus. co-incubated with antibodies or serums, infected with HOS cells expressing CD4 receptor and a coreceptor CCR5 / CXCR4. After 3 days of culture, the cells were washed with PBS, and the cells were lysed. The luciferase content was detected, and the cell lines stably expressing HIV-1 pseudovirus were further established, and lots of pseudoviruses were obtained. This method can greatly reduce the variation caused by the difference of host cells (PBMCs) in traditional methods and is a reproducible and promising in vitro neutralization method.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2006
【分類號】：R392

【參考文獻】

相關(guān)期刊論文 前1條

1 蘇玲,邢輝,羊海濤,羅小光,邵一鳴;中國首例人類免疫缺陷病毒(HIV-1)A亞型毒株的鑒定[J];病毒學(xué)報;1997年03期

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本文編號：1693039