本文關(guān)鍵詞：慢性難愈創(chuàng)面細(xì)菌生物膜體外模型的建立及分析　出處：《內(nèi)蒙古大學(xué)》2012年碩士論文　論文類(lèi)型：學(xué)位論文

  更多相關(guān)文章： 細(xì)菌生物膜 激光掃描共聚焦顯微鏡 免疫熒光技術(shù)

【摘要】：目的：建立慢性難愈創(chuàng)面臨床常見(jiàn)細(xì)菌的生物膜體外模型,利用激光共聚焦顯微鏡觀察細(xì)菌生物膜形成過(guò)程,探索細(xì)菌生物膜檢測(cè)的有效手段。方法：(1)建立細(xì)菌生物膜體外模型,摸索生物膜體外模型的建立條件,首先將傳統(tǒng)的平板培養(yǎng)法與引導(dǎo)片培養(yǎng)法進(jìn)行比較,在此基礎(chǔ)上,以引導(dǎo)片法體外培養(yǎng)生物膜,觀察不同時(shí)間點(diǎn)和溫度對(duì)生物膜形成的影響。(2)激光共聚焦顯微鏡觀察各菌株生物膜形成過(guò)程。采用FITC-ConA和碘化丙啶(PI)分別標(biāo)記多糖和細(xì)菌DNA,激光掃描共聚焦顯微鏡觀察生物膜形成情況。結(jié)果：(1)傳統(tǒng)的平板培養(yǎng)法最終測(cè)得的結(jié)果穩(wěn)定性較差,引導(dǎo)片培養(yǎng)法最終所得的結(jié)果較穩(wěn)定,試驗(yàn)重復(fù)性較好,37℃時(shí)生物膜的生長(zhǎng)狀態(tài)較22℃更好。(2)CLSM觀察可見(jiàn),培養(yǎng)24h時(shí),各菌株均有散在的綠色熒光,不密集。48h時(shí)綠色熒光明顯增多,部分與紅色熒光重疊,形成視野中黃色熒光,金黃色葡萄球菌明顯。在綠色熒光的強(qiáng)度和分布上,臨床菌株明顯多于標(biāo)準(zhǔn)株。培養(yǎng)72h時(shí)銅綠假單胞菌和其標(biāo)準(zhǔn)株的綠色熒光增多。培養(yǎng)5d時(shí)綠色熒光較分散。金黃色葡萄球菌在48h形成成熟的生物膜,銅綠假單胞菌則在72h。結(jié)論：(1)慢性創(chuàng)面臨床常見(jiàn)的金黃色葡萄球菌和銅綠假單胞菌均能形成生物膜。(2)采用免疫熒光技術(shù)和激光掃描共聚焦顯微鏡研究生物膜形成過(guò)程是一種簡(jiǎn)便可行的方法。金黃色葡萄球菌、銅綠假單胞菌形成生物膜的時(shí)間、生物膜的形態(tài)與特點(diǎn)都有顯著的差異。
[Abstract]:Objective: to establish a biofilm in vitro model of chronic refractory wounds clinical common bacteria, confocal microscopy observation of bacterial biofilm formation by laser, explore the effective means of bacterial biofilm detection. Methods: (1) establish the bacterial biofilm model in vitro, explore the conditions of establishment of biofilm model in vitro, the traditional plate culture method was compared with the guide piece culture method, on this basis, in order to guide plate biofilm method in vitro to observe the effect of different time and temperature on the biofilm formation. (2) the biofilm formation of each strain was observed by laser confocal microscope. FITC-ConA and propidium iodide (PI) were used to mark polysaccharide and bacterial DNA, and the formation of biofilm was observed by laser scanning confocal microscope. Results: (1) the results of the traditional plate culture method were not stable. The results obtained by the guided culture method were stable and the reproducibility was good. The growth state of biofilms was better than 22 degrees at 37 degrees. (2) CLSM observation showed that when the 24h was cultured, all the strains had the green fluorescence, which was not dense. At 48h, the green fluorescence increased obviously, partly overlapped with red fluorescence, forming yellow fluorescence in the field of vision, and the Staphylococcus aureus was obvious. In the intensity and distribution of green fluorescence, the clinical strains were obviously more than the standard strains. The green fluorescence of Pseudomonas aeruginosa and its standard strains were increased when 72h was cultured. When the 5D was cultured, the green fluorescence was more dispersed. Staphylococcus aureus forms a mature biofilm in 48h, and Pseudomonas aeruginosa is in 72h. Conclusions: (1) the common Staphylococcus aureus and Pseudomonas aeruginosa in the chronic wound surface can form the biofilm. (2) it is a simple and feasible method to study the process of biofilm formation by using immunofluorescence and laser scanning confocal microscopy. The time of the biofilm formation of Staphylococcus aureus and Pseudomonas aeruginosa, the morphological and characteristics of the biofilm are significantly different.
【學(xué)位授予單位】：內(nèi)蒙古大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R641;R-332

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相關(guān)期刊論文 前2條

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本文編號(hào)：1344979