本文關鍵詞：恥垢分枝桿菌與結(jié)核分枝桿菌抗原的交叉免疫反應研究　出處：《重慶醫(yī)科大學》2017年碩士論文　論文類型：學位論文

  更多相關文章： 結(jié)核分枝桿菌 恥垢分枝桿菌 交叉抗原 免疫應答

【摘要】：目的探討恥垢分枝桿菌(Mycobacteria smegmatis,MS)與結(jié)核分枝桿菌(Mycobacteria tuberculosis,MTB)共同抗原及其免疫原性,為恥垢分枝桿菌作為結(jié)核潛在疫苗的研究提供科學依據(jù)。方法以低溫超聲破碎、活菌碾磨破碎兩種方法提取MS和MTB菌體抗原,以及自己收集的培養(yǎng)后期上清濾液蛋白,通過Western blot觀察菌體抗原與抗血清的免疫交叉反應。將MS超聲破碎抗原和MTB超聲破碎抗原分別與不完全弗氏佐劑混勻后肌肉注射于小鼠,免疫不同次數(shù)后7天,以Elispot技術檢測小鼠脾臟淋巴細胞經(jīng)不同細菌抗原分別刺激后能產(chǎn)生抗原特異性IFN-γ的淋巴細胞水平,流式細胞學技術檢測各類細胞因子水平,ELISA技術檢測血清中抗體效價水平。結(jié)果Western blot結(jié)果顯示MS與MTB間存在一些主要分布在10-100KD的共同抗原,能與兩細菌的抗血清產(chǎn)生較強的交叉免疫反應;ELISPOT及流式細胞學結(jié)果顯示經(jīng)兩細菌分別致敏的小鼠均能兩種分枝桿菌抗原產(chǎn)生以細胞免疫為主的免疫應答,隨免疫次數(shù)的增加免疫反應也增強,免疫兩針接近高峰,免疫三針達到平衡并出現(xiàn)機體免疫系統(tǒng)的平衡調(diào)節(jié)。CD4+T及CD8+T淋巴細胞分泌IL-2、IFN-γ水平均明顯高于未免疫組(P0.05),且對非分枝桿菌屬的布氏桿菌對照抗原幾乎不產(chǎn)生細胞免疫應答。ELISA結(jié)果顯示兩細菌抗原分別免疫的小鼠血清中含有大量能與共同抗原特異性結(jié)合的抗體,并且隨免疫次數(shù)的增加而增高,免疫三針接近高峰。結(jié)論MS與MTB間存在這些共同抗原使MS致敏動物能引起針對結(jié)核分枝桿菌特異性免疫反應,為MS用于結(jié)核新疫苗佐劑等研究提供科學依據(jù)。
[Abstract]:Objective to investigate the Mycobacterium smegmatis (Mycobacteria smegmatis, MS) and Mycobacterium tuberculosis (Mycobacteria tuberculosis MTB) common antigen and its immunogenicity as Mycobacterium smegmatis as potential vaccine of tuberculosis and provide scientific basis. Methods with low temperature ultrasonic crushing, milling and extraction of MS bacteria MTB antigen broken two methods. And his collection of late culture filtrate proteins, observe the immune cross reaction antigen and antiserum by Western blot. The MS ultrasonic antigen and MTB supersonic antigen respectively with incomplete Freund's adjuvant mixed intramuscular injection in mice 7 days after immunization, at different times, in order to detect the spleen lymphocytes of mice by different Elispot technology bacterial antigens were stimulated to produce antigen-specific IFN- lymphocyte gamma level, flow cytometry detection of cytokines level of ELISA. Antibody titers were detected in serum. The results showed that some Western blot mainly distributed in the common antigen of the existence of 10-100KD between MTB and MS, can produce strong cross immune response and antibody two bacteria; ELISPOT and flow cytometry analysis showed that the two bacteria were sensitized mice to the immune response to cell immunity is the main two kinds of mycobacterial antigens, the immune response increased with immunization times also enhanced immune, two needle close to the peak, three needle balance and immune immune system regulates the balance of.CD4+T and CD8+T lymphocytes secreting IL-2, IFN- gamma average water was significantly higher than that in non immune group (P0.05), and almost no cellular immune antigen the results showed that serum.ELISA response in mice of two bacterial antigen, contains a large number can be combined with the common antigen of Brucella non Mycobacterium control The antibody increased with the increase of immunization times. The three immunization needles approached the peak. Conclusion these common antigens exist between MS and MTB, so that MS sensitized animals can cause specific immune responses against Mycobacterium tuberculosis, and provide scientific basis for MS in the research of new vaccine adjuvant for tuberculosis.

【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R392

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