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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 朱士維 | |
| dc.contributor.author | Chen-Li Lin | en |
| dc.contributor.author | 林晨立 | zh_TW |
| dc.date.accessioned | 2021-06-16T16:44:13Z | - |
| dc.date.available | 2015-08-28 | |
| dc.date.copyright | 2012-08-28 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-08-21 | |
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Investugate the correlation between collagen fiber and photonic band gap structure by second order susceptibility. Graduate Institute of Physics, National Taiwan University Master Thesis. (2011). [29] S.-W. Chu, S.-P. Tai, C.-K. Sun & C.-H. Lin. Selective Imaging in Second-Harmonic-Generation Microscopy by Polarization Manipulation. Applied Physics Letters. 91, 103903 (2007). [30] K. Tamura, T Shimada, E. Ono, Y. Tanaka, A. Nagatani & S.-I Higashi. Why green fluorescent proteins have not been observed in the vacuoles of higher plants. The Plant Journal. 35, 545-555 (2003). [31] S. V. Plotnikov, A. C. Millard, P. J. Campagnola & W. A. Mohler. Characterization of the myosin-based source for second harmonic generation from muscle sarcomeres. Biophysical Journal. 90, 328–339 (2006). [32] J.-Y. Yu. A subcellular-resolution spectro-ophthalmoscope. Graduate Institute of Physics, National Taiwan University Master Thesis. (2008). [33] Spectra-Physics, Inc. Mai Tai–Series, High-Performance, Mode-Locked, Ti: Sapphire Laser, User’s Manual. Spectra-Physics, Inc. (2007). [34] D. Semwogerere & E. R. Weeks. Confocal Microscopy. Encyclopedia of Biomaterials and Biomedical Engineering. (2005). [35] B. Martin. Second harmonic imaging of intrinsic signals in muscle fibers in situ. Journal of Biomedical Optics. 9(5), 882–892 (2004). [36] G. A. Blab. Two-photon excitation action cross-sections of the autofluorescent proteins. Chemical Physics Letters. 350, 71-77 (2001). | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/63475 | - |
| dc.description.abstract | 研究神經科學的重要性已與日俱增，這個領域最具吸引力的生物組織研究就是大腦的研究，在此一環最重要的模式動物之一就是果蠅幼蟲。腦的研究之所以重要不僅是其具備其他組織上能表現的接收、傳遞與反應外，最重要的是具備了額外的思考機制。雖然在許多傑出的研究成果顯示大腦神經網路地圖已被基因工程學完整的標定出來，但這樣的結果只顯示了神經網路系統形式上的連結，而非真正功能上的連結。為了更進一步去證明兩者連結關係的關聯性，我們將利用鈣離子訊號染劑 (calcium dye) 與電位訊號染劑 (voltage sensitive dye) 配合共焦顯微技術 (confocal microscopy) 做為我們量測神經訊號的工具，然而這些染劑都必須透過二甲基亞碸 (DMSO) 此有機溶劑幫助溶解，因此我們希望透過非線性光學技術來檢驗DMSO是否會對果蠅幼蟲樣本的分子結構造成影響。根據二倍頻 (second harmonic generation-SHG) 信號產生必須與材料的分子結構有密切關係，因此二倍頻信號強弱變化將被分子結構的變化影響。我們分析了浸泡在等滲透壓的生理緩衝液 (PBS) 與DMSO中的樣本，發現浸泡DMSO樣本的二倍頻信號將明顯增強，這暗示樣本的分子結構已經在不同溶液下產生改變。我們也另外發現浸泡DMSO會導致基因轉殖的綠螢光蛋白 (green fluorescence protein-GFP) 在近紅外光激發下，雙光子螢光 (two-photon fluorescence) 信號有消失的跡象。相信這些結果對未來以果蠅幼蟲做為研究神經科學樣本的學者們提供了在細胞染色上值得注意的方向，也讓基因轉殖工程多了一份參考資料。 | zh_TW |
| dc.description.abstract | Neuron science research has been more and more popular recently. The most attractive organism in this study is brain, and one of the most famous model samples is drosophila larva. Although almost whole brain structure map studied by genetic engineering has been constructed, it still only provides structural connection, but not functional connection. In order to further prove the correlation of these two connections, we use calcium dye and voltage sensitive dye as our neuron signal indicator and construct a confocal microscopy to detect it. Both of these dyes should be dissolved into dimethyl sulfoxide (DMSO) solution, thus, analyzing the influence of DMSO on our sample drosophila larva is also important. According to the principle of second harmonic microscopy (SHG), signal intensity will depend on the molecular structure of samples. We analyze the SHG signal of our sample after soaking in phosphate buffered saline (PBS) solution and DMSO solution respectively. Samples soaked in DMSO lead to the enhancement of SHG signal, and that implies there would be some structural changes in our organism samples. We also find that the performance of green fluorescence protein (GFP) of two-photon fluorescence (TPF) will vanish after DMSO soaked. This result may provide a new perspective for those who want to load dyes on drosophila larvae. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T16:44:13Z (GMT). No. of bitstreams: 1 ntu-101-R98245015-1.pdf: 15871931 bytes, checksum: b110041b1cccf07b38efc4b4f286efb7 (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | 目錄
口試委員審定書 誌謝…………………………………………………………………………... I 中文摘要……………………………………………………………………... III 英文摘要……………………………………………………………………... IV 目錄……………………………………………………………………........... V 圖目錄……………………………………………………………………....... VII 表目錄……………………………………………………………………....... IX 第一章 導論 1 1-1 顯微鏡的發展……………………………………………………… 1 1-2 顯微鏡與生物醫學………………………………………………… 1 1-3 非線性光學………………………………………………………… 2 1-4 光學與神經科學的結合…………………………………………… 3 1-5 此論文研究與價值………………………………………………… 4 第二章 基本原理 5 2-1 二階非線性光學─二倍頻 (SHG) ……………………………...… 5 2-2 倍頻訊號與分子密度的關係……………………………………… 8 2-3 螢光與雙光子螢光 Two-photon Fluorescence (TPF) …………10 第三章 實驗架設與樣本處理 12 3-1顯微鏡系統…………………………………………………………. 12 3-1-1 Olympus FV300 共焦顯微鏡系統………………………… 13 3-1-2 信號來源驗證系統………………………………………… 14 3-1-3 Olympus FV1000 MPE 多光子共焦顯微系統……………. 15 3-2 樣本準備 16 3-2-1神經單細胞 (cortical neuronal cell) ………………………. 17 3-2-2 果蠅幼蟲 (drosophila larva) ……………………………… 17 3-2-2(1) 單光子鈣離子信號實驗的樣本準備…………………… 17 3-2-2(2) 二倍頻與雙光子實驗的樣本準備……………………… 18 第四章 結果分析 19 4-1 神經單細胞在鈣離子染劑下的螢光表現………………………… 19 4-2 鈣離子染劑在表現綠螢光蛋白(GFP)轉植果蠅幼蟲的螢光表現.. 22 4-3 二甲基亞碸對果蠅幼蟲的二倍頻信號的影響…………………… 24 4-4 二甲基亞碸對果蠅幼蟲的雙光子信號的影響…………………… 31 第五章 結論 34 參考文獻 35 圖目錄 圖(2-1-1) 二倍頻激發機制：分子被兩顆光子 2(ℏω) 激發，轉而釋放 ℏ(2ω)=ℏω_SHG………………………………………………………………………………………………………………..6 圖(2-3-1) 單光子螢光激發機制：螢光分子吸收單一顆光子能量，轉而放出螢光的能階躍遷………………………………………………………………………………………………………….10 圖(2-3-2) 雙光子螢光激發機制：螢光分子同時吸收兩顆光子能量，轉而放出螢光的能階躍遷………………………………………………………………………………………………………11 圖(3-1-1)系統概略圖………………………………………………………………………………………….12 圖(3-1-2) 473nm 藍光雷射(laser)，通過掃描系統(scanning system)，經物鏡(Ob.)聚焦於樣本(specimen)，再由反射頻道收集信號……………………………………………………….13 圖(3-1-3) 1040nm 近紅外線雷射(laser)，通過掃描系統(scanning system)，經物鏡(Ob.)聚焦於樣本(specimen)，再由穿透頻道收集信號………………………………………………..14 圖(3-1-4) 970nm 近紅外線雷射(laser)，通過掃描系統(scanning system)，經物鏡(Ob.)聚焦於樣本(specimen)，再由穿透頻道收集二倍頻與雙光子螢光信號……………..….15 圖(4-1-1) 為單細胞的神經細胞 (cortical neuronal cell) 在鈣離子染劑 (Fluo2) 的附著下取得的螢光影像… ………………………………………………………………………………….…19 圖(4-1-2) 左圖是在未滴入高鉀離子溶液激發神經訊號的影像強度，右圖在離子溶液激發的瞬間整體出現了亮度改變，尤其在細胞與細胞間的絲狀連結…………..….20 圖(4-1-3) 三個圈取區域在高鉀離子溶液滴入（時間 29.48 s）後造成神經訊號激發前後的螢光信號強弱變化圖……………………………………………………………...……………..20 圖(4-1-4) 三個圈選細胞受高鉀離子水溶液滴入（時間 29.48 s）後造成神經訊號激發後於各不同時間產生的螢光強度隨時間變化…………………………………………..……..21 圖(4-2-1) 鈣離子染劑附著在表現GFP果蠅幼蟲神經細胞的螢光影像…………...……22 圖(4-2-2) 果蠅幼蟲樣本被離子溶液滴下同時造成位移現象…………………………....…..23 圖(4-2-3) 標定的神經細胞在抽動後回到原處，並分析此動作前後的螢光信號強弱 …………………………………………………………………………………………………………………....…..24 圖(4-3-1) 果蠅幼蟲全身肌肉的二倍頻影像，黑色框框是每次實驗進行影像分析的第三節爬行體節。右邊顏色條表示信號強弱範圍…………………………………………..….25 圖(4-3-2) 幼蟲肌肉在浸泡二甲基亞碸前後的二倍頻影像，data1 選定分析的肌肉是在整張影像中最亮的肌肉。Data2 選定的是同一條肌肉……………………….…………...27 圖(4-3-3) 圖(4-3-2) 放大3倍的肌肉影像。在兩組資料中各選出兩條指向性相近的肌肉橫紋做分析………………………………………………………………………………………………..….27 圖(4-3-4) 肌肉橫紋在浸泡PBS與DMSO的空間出現頻率.…………………………..………29 圖(4-4-1) 果蠅幼蟲的全身雙光子螢光影像………………………………………………………...31 圖(4-4-2) GFP的激發頻譜…………………………………………………………………………….……32 圖(4-4-3) 果蠅幼蟲第三體節運動肌肉的Z軸雙光子螢光影像，數字表示影像距離幼蟲表皮的深度，單位：μm…………………………………………………………………………….33 表目錄 表(3-1-1) 三系統使用的光源、物鏡、濾波片、收集的信號及收光頻道，系統一、 二為單頻道收光系統，系統三為雙頻道收光系統………………………………………….…….12 表(4-1-1) 三個圈取區域在高鉀離子溶液激發前後的螢光信號強弱變化數值表 …………………………………………………………………………………………………………………….…..21 表(4-3-1) 兩組實驗資料浸泡在不同溶液之下的二倍頻影像強度整理……………….…28 | |
| dc.language.iso | zh-TW | |
| dc.title | 以多光子顯微技術分析二甲基亞碸對果蠅幼蟲的影響 | zh_TW |
| dc.title | The Effect of DMSO in Drosophila Larvae Studied by Multiphoton Microscopy | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 江安世,林彥穎,潘建源 | |
| dc.subject.keyword | 共焦顯微鏡,鈣離子染劑,神經電位染劑,二甲基亞碸,二倍頻,綠螢光蛋白,雙光子螢光, | zh_TW |
| dc.subject.keyword | confocal microscopy,calcium dye,voltage sensitive dye,dimethyl sulfoxide,second harmonic microscopy,phosphate buffered saline,green fluorescence protein,two-photon fluorescence, | en |
| dc.relation.page | 37 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-08-21 | |
| dc.contributor.author-college | 理學院 | zh_TW |
| dc.contributor.author-dept | 應用物理所 | zh_TW |
| 顯示於系所單位： | 應用物理研究所 | |
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