發(fā)布時(shí)間：2018-01-14 16:42

  本文關(guān)鍵詞：日本血吸蟲細(xì)胞培養(yǎng)條件的篩選和抗原含量動(dòng)態(tài)變化的研究　出處：《武漢大學(xué)》2005年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 日本血吸蟲 細(xì)胞培養(yǎng)條件 正交試驗(yàn) 琥珀酸脫氫酶 乳酸脫氫酶 葡萄糖-6-磷酸脫氫酶 細(xì)胞外基質(zhì) 免疫細(xì)胞化學(xué) 抗原

【摘要】：本文以琥珀酸脫氫酶(SDH)、乳酸脫氫酶(LDH)和葡萄糖-6-磷酸脫氫酶(G-6-PDH)為評(píng)價(jià)指標(biāo),運(yùn)用正交試驗(yàn)法篩選日本血吸蟲細(xì)胞的培養(yǎng)條件,探討這3個(gè)評(píng)價(jià)指標(biāo)的可靠性;并用篩選出的培養(yǎng)條件培養(yǎng)日本血吸蟲細(xì)胞,研究日本血吸蟲培養(yǎng)細(xì)胞抗原含量的動(dòng)態(tài)變化。所獲結(jié)果簡(jiǎn)述如下: 1.日本血吸蟲細(xì)胞培養(yǎng)條件的選擇 以SDH、LDH和G-6-PDH為評(píng)價(jià)指標(biāo),運(yùn)用正交試驗(yàn)法研究合成培養(yǎng)基(TC-199、RPMI-1640和DMEM)、細(xì)胞外基質(zhì)(鼠尾膠、肝基質(zhì)與肺基質(zhì))和血清濃度(5%、10%及20%)3個(gè)因素在3個(gè)不同水平對(duì)蟲齡為21d的日本血吸蟲培養(yǎng)細(xì)胞的影響。培養(yǎng)第5d,對(duì)日本血吸蟲培養(yǎng)細(xì)胞進(jìn)行SDH染色;培養(yǎng)第14d,分別進(jìn)行LDH、G-6-PDH染色;培養(yǎng)第28d,進(jìn)行LDH染色。將染色后的培養(yǎng)細(xì)胞置于Olympus-BH_2顯微鏡下觀察并拍照,HPIAS-2000圖像分析儀測(cè)量它們的含量,用SPSS10.0作統(tǒng)計(jì)分析。結(jié)果顯示:培養(yǎng)早期,不論以SDH或LDH,還是G-6-PDH為指標(biāo),RPMI-1640含20%小牛血清組成的培養(yǎng)基培養(yǎng)接種于肝基質(zhì)上的日本血吸蟲細(xì)胞,培養(yǎng)細(xì)胞的SDH、LDH、G-6-PDH活性最強(qiáng)。血清濃度對(duì)培養(yǎng)細(xì)胞酶活性的影響最大,其次是細(xì)胞外基質(zhì),影響最小的是合成培養(yǎng)基。其中,血清濃度對(duì)3種酶活性的影響差異均顯著(P0.01或P0.05);基質(zhì)對(duì)SDH、LDH活性的影響差異顯著(P0.01或P0.05),而對(duì)G-6-PDH活性的影響差異不顯著(P0.05);合成培養(yǎng)基的影響均無差異(P0.05)。培養(yǎng)28d以后,DMEM含5%小牛血清組成的培養(yǎng)基培養(yǎng)接種于肝基質(zhì)上的日本血吸蟲細(xì)胞,培養(yǎng)細(xì)胞的LDH活性最強(qiáng)。血清濃度對(duì)培養(yǎng)細(xì)胞LDH活性的影響最大,其次是細(xì)胞外基質(zhì),影響最小的是合成培養(yǎng)基。血清濃度和細(xì)胞外基質(zhì)對(duì)LDH活性的影響差異均顯著(P0.05),合成培養(yǎng)基的影響均無顯著性差異(P0.05)。因此,選擇3種酶中任一,均可用來評(píng)價(jià)日本血吸蟲細(xì)胞培養(yǎng)條件的優(yōu)劣。在培養(yǎng)早期,選擇三種合成培養(yǎng)基中的任一種與20%小牛血清組成的培養(yǎng)基培養(yǎng)接種于肝基質(zhì)上的日本血吸蟲細(xì)胞,培養(yǎng)細(xì)胞的SDH、LDH、G-6-PDH活性最強(qiáng)。培養(yǎng)28d以后,三種培
[Abstract]:In this paper, succinate dehydrogenase (SDH), lactate dehydrogenase (LDH) and glucose -6- phosphate dehydrogenase (G-6-PDH) as the evaluation index, screening of Schistosoma japonicum cell culture conditions by using orthogonal test method, the reliability of the 3 indexes; and screened cultured cells of Schistosoma japonicum and Schistosoma japonicum dynamic changes of cell culture antigen content. The results are summarized as follows:
Selection of cell culture conditions for Schistosoma japonicum 1.
In SDH, LDH and G-6-PDH as the evaluation index, using orthogonal test method based on Synthesis of culture (TC-199, RPMI-1640 and DMEM), extracellular matrix (rat tail collagen, liver and lung stromal matrix) and serum concentration (5%, 10% and 20%) 3 factors at 3 different levels of instar 21d the schistosome effects in cultured cells cultured in 5D cells. SDH staining, cultivation of Schistosoma japonicum; cultured in 14d, were analyzed by LDH and G-6-PDH staining; cultured 28d, LDH staining. After staining the cells cultured in Olympus-BH_2 observed and photographed under the microscope, measuring their content of HPIAS-2000 image analyzer, statistical analysis by using SPSS10.0. The results showed that early training, either at SDH or LDH, or G-6-PDH as the index, RPMI-1640 containing 20% calf serum. The cultured in culture medium on liver stromal cells of Schistosoma japonicum, the culture cells of SDH, LDH, G-6-PDH activity was the strongest. Effect of serum concentration on enzyme activity in cultured cells, followed by extracellular matrix, the impact is minimal synthetic medium. The difference of serum concentration of 3 enzymes were significantly (P0.01 or P0.05); matrix of SDH, influence the activity of LDH significantly (P0.01 or P0.05), but not significant effect on the activity of G-6-PDH difference (P0.05); there was no significant difference in effects of synthetic medium (P0.05). After 28d culture, DMEM containing 5% calf serum. The cultured in culture medium on liver stromal cells of Schistosoma japonicum, cultured cells LDH activity. The effects of serum concentration on LDH activity in cultured cells the largest, followed by the extracellular matrix, the impact is minimal synthetic medium. The serum concentration of extracellular matrix and the influence on the activity of LDH were significantly different (P0.05), there were no significant differences between the effects of synthetic medium (P0.05). Therefore, 3 kinds of enzyme in any, Can be used to evaluate the culture conditions of cells from Schistosoma japonicum. In the early stage of culture, culture medium were inoculated in the liver matrix choose three synthetic medium one and 20% calf serum. The cultured cells of Schistosoma japonicum, SDH, LDH, the strong G-6-PDH activity. 28d culture after three.

【學(xué)位授予單位】：武漢大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類號(hào)】：R383

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