雙光子掃描結構照明顯微術

以作者查詢圖書館館藏

、以作者查詢臺灣博碩士

、以作者查詢全國書目

、勘誤回報

、線上人數：17

、訪客IP：3.144.163.15

姓名

葉佳樺(Chia-Hua Yeh) 查詢紙本館藏

畢業系所

光電科學與工程學系

論文名稱

雙光子掃描結構照明顯微術
(Two-photon Scanning Structured Illumination Microscopy)

相關論文

★ 非反掃描式平行接收之雙光子螢光超光譜顯微術	★ 以二次通過成像量測架構及降低誤差迭代演算法重建人眼之點擴散函數
★ LASER光源暨LED在老鼠毛生長的低能量光治療比較分析	★ 應用線狀結構照明提升雙光子顯微鏡解析度
★ 以同調結構照明顯微術進行散射樣本解析度之提升	★ 掃描式二倍頻結構照明顯微術
★ 小貓自泵相位共軛鏡於數位光學相位共軛與時間微分之研究	★ 鏡像輔助斷層掃描相位顯微鏡
★ 以數位全像術重建多波長環狀光束之研究	★ 相位共軛反射鏡用於散射介質中光學聚焦之研究
★ 雙光子螢光超光譜顯微術於多螢光生物樣本之研究	★ 倍頻非螢光基態耗損超解析之顯微成像方法
★ 葉綠素雙光子螢光超光譜影像於光合作用研究之應用	★ 微投影光學切片超光譜顯微術
★ 使用結構照明顯微術觀察活體小鼠毛囊生長週期之變化	★ 一次性多角度漫射光譜量測系統

檔案

[Endnote RIS 格式]

[Bibtex 格式]

[相關文章]

[文章引用]

[完整記錄]

[館藏目錄]

[檢視]

[下載]

本電子論文使用權限為同意立即開放。
已達開放權限電子全文僅授權使用者為學術研究之目的，進行個人非營利性質之檢索、閱讀、列印。
請遵守中華民國著作權法之相關規定，切勿任意重製、散佈、改作、轉貼、播送，以免觸法。

摘要(中)

雙光子螢光顯微術比起傳統的廣域顯微鏡有著高縱向解析度的特點和高穿透深度的優點，故廣泛地被應用在生物醫學研究與臨床實驗。然而雙光子顯微鏡的空間解析度受限於光學繞射極限，低於繞射極限結構的樣本將無法被解析。為了在生物厚組織中也能取得優秀於傳統雙光子顯微術解析度的影像，本研究將結合結構照明與雙光子顯微術的概念開發一套超解析度影像系統，稱之為雙光子掃描結構照明顯微術(Two-photon scanning structured illumination microscopy, TPS-SIM)。
在此系統中，強聚焦雷射光激發樣本並產生雙光子螢光訊號，在雷射掃描過程中藉由二維相機收集每個激發光點的訊號並組成影像，若控制其光點掃描路徑使激發光強度分布類似方波，在此結構照明下將可獲得傳統影像系統無法擁有的高空間頻率資訊。多筆具有結構照明的影像資料經過演算重建後，將可以產生更高解析度的影像。在本論文中將會詳述系統成像之理論模型及影像重建之演算法，並討論條紋週期大小、佔空比(Duty cycle)、等效明晰度與影像訊雜比對解析度的影響。藉由測量螢光奈米小球的螢光強度分布，可知TPS-SIM在橫向上最大解析度提升倍率為1.42倍。結合雙光子系統所具有的光學切片能力，本系統可在蘇木素•伊紅染色的動物組織切片與老鼠皮膚中取得三維影像，除橫向解析度的提升外，其縱向解析度最大可提升至1.67倍。

摘要(英)

Compared with conventional wide-field microscopy, two-photon microscopy (TPM) has advantages of inherent axial resolution and high penetration. Therefore, TPM has been widely applied to bio-medical and clinical researches. However, the spatial resolution of TPM is restricted by the optical diffraction limit so structures smaller than the limit can’t be resolved. To improve the resolution of TPM image in depth tissue, this research will integrate the concept of structured illumination and TPM to develop an imaging system called two-photon scanning structured illumination microscopy (TPS-SIM).
In this system, laser beam is tightly focused onto sample to excite two-photon fluorescence signals. The excited signals are imaged and integrated by 2D camera point by point to form an image. During the scanning procedure, the path of the excitation spot is modulated to form an effective structured illumination with a square-wave intensity distribution. Under this structure illumination, higher spatial frequency out of the reach of the conventional wide field microscopy can thus be obtained. An image with improved resolution can be reconstructed though multiple patterned images with different phases. In this research, the theory of the image formation and the image reconstruction algorithm will be clearly introduced. The effects that the period, duty cycle, effective modulation depth of pattern and SNR (signal and noise ratio) may have on the resolution improvement will be discussed. By measuring the fluorescence intensity distribution of the nanoparticles, the maximum resolution improvement ratio of TPS-SIM is around 1.42-fold. Combined the optical sectioning ability of two-photon excitation, 3D images can be obtained in H&E stained sectioned bio-tissues and fluorescence stained whole-mounted mouse skin. In addition to improvement in lateral resolution, the maximum improvement ratio in axial is around 1.67-fold.

關鍵字(中)

★ 非線性顯微術
★ 螢光顯微術
★ 超解析顯微術
★ 結構照明
★ 切片影像

關鍵字(英)

★ Nonlinear microscopy
★ Fluorescence microscopy
★ Super-resolution microscopy
★ Structured illumination
★ Sectioning image

論文目次

摘要 ii
Abstract iv
目錄 v
圖索引 vi
第一章緒論 1
1.1 研究背景 1
1.1.1 光學顯微系統 1
1.1.2 光學切片顯微術 5
1.1.3 超解析顯微術 6
1.2 研究目的與動機 9
第二章原理 11
2.1 雙光子螢光顯微術 11
2.2 成像原理 13
2.3 廣域結構照明螢光顯微術 15
2.4 雙光子掃描結構照明顯微術 20
第三章系統架構 24
3.1 實驗架構 24
3.2 實驗參數 27
第四章數據模擬與結果 31
4.1 實驗模擬 31
4.2 樣本與雜訊模擬 34
4.2.1模擬樣本 35
4.2.2雜訊模型 36
4.2.3 影像重建演算法與結果 37
4.2.4 模擬結果 40
第五章實驗結果 50
5.1 奈米螢光小球影像 50
5.2 H&E stained組織切片樣本量測 52
5.2.1 TPS-SIM影像 52
5.2.2 居留條紋處理 56
5.2.3 調整Duty cycle 58
5.3 三維TPS-SIM影像 60
第六章結論 64
參考文獻 66

參考文獻

[1] V. H. Albert, The Origins of the Telescope, History of Science and Scholarship in the Netherlands (Amsterdam University Press, Netherlands, 2011).
[2] L. Jardine, The Curious Life of Robert Hooke: The Man Who Measured London (New York: Harper Collins Publishers, New York, 2005).
[3] L. Day, Biographical Dictionary of the History of Technology (Taylor & Francis, Francis, 2003).
[4] E. Abbe, "Beiträge zur theorie des mikroskops und der mikroskopischen wahrnehmung," Archiv für Mikroskopische Anatomie 9, 413-468 (1873).
[5] J. W. Goodman, Introduction to Fourier Optics, 3rd ed. ( Roberts & Company Publishers, United States, 2005).
[6] M. W. Davidson and M. Abramowitz, "Optical Microscopy," in Encyclopedia of Imaging Science and Technology (John Wiley & Sons, Inc., 2002).
[7] F. Helmchen and W. Denk, "Deep tissue two-photon microscopy," Nat Methods 3, 235-235 (2006).
[8] T. Horio and H. Hotani, "Visualization of the dynamic instability of individual microtubules by dark-field microscopy," Nature 321, 605-607 (1986).
[9] C. R. Burch and J. P. P. Stock, "Phase-Contrast Microscopy," Journal of Scientific Instruments 19, 71 (1942).
[10] G. Popescu, T. Ikeda, R. R. Dasari, and M. S. Feld, "Diffraction phase microscopy for quantifying cell structure and dynamics," Optics Letters 31, 775-777 (2006).
[11] F. W. D. Rost, Fluorescence Microscopy, 1st ed. (Cambridge University Press, New York, 1992).
[12] J. S. Ploem, "The use of a vertical illuminator with interchangeable dichroic mirrors for fluorescence microscopy with incidental light," Zeitschrift fur wissenschaftliche Mikroskopie und mikroskopische Technik 68, 129-142 (1967).
[13] M. A. Bopp, G. Tarrach, M. A. Lieb, and A. J. Meixner, "Super-resolution fluorescence imaging of single dye molecules in thin polymer films," Journal of Vacuum Science & Technology A 15, 1423-1426 (1997).
[14] C. R. Parish, "Fluorescent dyes for lymphocyte migration and proliferation studies," Immunology and cell biology 77, 499-508 (1999).
[15] L. Wu and K. Burgess, "Fluorescent amino- and thiopyronin dyes," Organic letters 10, 1779-1782 (2008).
[16] E. J. Wood, Molecular probes: Handbook of fluorescent probes and research chemicals (Molecular Probes Inc, US, 1994).
[17] M. Chalfie, Y. Tu, G. Euskirchen, W. W. Ward, and D. C. Prasher, "Green fluorescent protein as a marker for gene expression," Science 263, 802-805 (1994).
[18] G.-J. Kremers, S. G. Gilbert, P. J. Cranfill, M. W. Davidson, and D. W. Piston, "Fluorescent proteins at a glance," J Cell Sci 124, 157-160 (2010).
[19] A. Miyawaki, J. Llopis, R. Heim, J. M. McCaffery, J. A. Adams, M. Ikura, and R. Y. Tsien, "Fluorescent indicators for Ca2+ based on green fluorescent proteins and calmodulin," Nature 388, 882-887 (1997).
[20] F. Yang, L. G. Moss, and G. N. Phillips, "The molecular structure of green fluorescent protein," Nat Biotech 14, 1246-1251 (1996).
[21] R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, "Resolution enhancement by subtraction of confocal signals taken at different pinhole sizes," Micron 34, 293-300 (2003).
[22] M. Von Tiedemann, A. Fridberger, M. Ulfendahl, and J. B. De Monvel, "Image adaptive point-spread function estimation and deconvolution for in vivo confocal microscopy," Microsc Res Techniq 69, 10-20 (2006).
[23] P. Vesely, Handbook of Biological Confocal Microscopy, 3rd ed. (Springer Science, New York, 2007).
[24] A. Weigel, D. Schild, and A. Zeug, "Resolution in the ApoTome and the confocal laser scanning microscope: comparison," J Biomed Opt 14, - (2009).
[25] T. Shimozawa, K. Yamagata, T. Kondo, S. Hayashi, A. Shitamukai, D. Konno, F. Matsuzaki, J. Takayama, S. Onami, H. Nakayama, Y. Kosugi, T. M. Watanabe, K. Fujita, and Y. Mimori-Kiyosue, "Improving spinning disk confocal microscopy by preventing pinhole cross-talk for intravital imaging," P Natl Acad Sci USA 110, 3399-3404 (2013).
[26] W. Denk, J. H. Strickler, and W. W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990).
[27] D. A. VanderSijs, U. A. VanderHeide, J. Sytsma, H. C. Gerritsen, and Y. K. L. Levine, "Confocal laser scanning fluorescence microscopy using two photon excitation for imaging muscle fibres," J Muscle Res Cell M 17, 120-120 (1996).
[28] C. J. R. Sheppard and M. Gu, "Image-Formation in 2-Photon Fluorescence Microscopy," Optik 86, 104-106 (1990).
[29] A. Diaspro and M. Robello, "Two-photon excitation of fluorescence for three-dimensional optical imaging of biological structures," J Photoch Photobio B 55, 1-8 (2000).
[30] P. Theer, M. T. Hasan, and W. Denk, "Two-photon imaging to a depth of 1000 mu m in living brains by use of a Ti : Al2O3 regenerative amplifier," Optics Letters 28, 1022-1024 (2003).
[31] W. R. Zipfel, R. M. Williams, and W. W. Webb, "Nonlinear magic: multiphoton microscopy in the biosciences," Nat Biotechnol 21, 1368-1376 (2003).
[32] F. Helmchen and W. Denk, "Deep tissue two-photon microscopy," Nat Methods 2, 932-940 (2005).
[33] K. Svoboda and R. Yasuda, "Principles of Two-Photon Excitation Microscopy and Its Applications to Neuroscience," Neuron 50, 823-839 (2006).
[34] C. R. Stoltzfus and A. Rebane, "High contrast two-photon imaging of fingermarks," Sci Rep 6, 24142 (2016).
[35] M. A. A. Neil, R. Juskaitis, and T. Wilson, "Method of obtaining optical sectioning by using structured light in a conventional microscope," Optics Letters 22, 1905-1907 (1997).
[36] M. A. A. Neil, Squire, R. Juškaitis, PI. Bastiaens, and T. Wilson, "Wide-field optically sectioning fluorescence microscopy with laser illumination," J. Microscopy 197, 1-4 (2000).
[37] J. Mitic, T. Anhut, M. Meier, M. Ducros, A. Serov, and T. Lasser, "Optical sectioning in wide-field microscopy obtained by dynamic structured light illumination and detection based on a smart pixel detector array," Optics Letters 28, 698-700 (2003).
[38] D. Karadaglic and T. Wilson, "Image formation in structured illumination wide-field fluorescence microscopy," Micron 39, 808-818 (2008).
[39] D. Karadaglic, "Image formation in conventional brightfield reflection microscopes with optical sectioning property via structured illumination," Micron 39, 302-310 (2008).
[40] K. Wicker and R. Heintzmann, "Single-shot optical sectioning using polarization-coded structured illumination," J Opt-Uk 12(2010).
[41] J. Qian, M. Lei, D. Dan, B. Yao, X. Zhou, Y. Yang, S. Yan, J. Min, and X. Yu, "Full-color structured illumination optical sectioning microscopy," Sci Rep 5, 14513 (2015).
[42] K. Nienhaus and G. U. Nienhaus, "Where Do We Stand with Super-Resolution Optical Microscopy?," J Mol Biol 428, 308-322 (2016).
[43] K. I. Mortensen, L. S. Churchman, J. A. Spudich, and H. Flyvbjerg, "Optimized localization analysis for single-molecule tracking and super-resolution microscopy," Nat Methods 7, 377-U359 (2010).
[44] M. Vega, J. Mateos, R. Molina, and A. K. Katsaggelos, "Super-Resolution of Multispectral Images," Comput J 52, 153-167 (2009).
[45] L. Schermelleh, R. Heintzmann, and H. Leonhardt, "A guide to super-resolution fluorescence microscopy," J Cell Biol 190, 165-175 (2010).
[46] S. W. Hell and J. Wichmann, "Breaking the Diffraction Resolution Limit by Stimulated-Emission-Stimulated-Emission-Depletion Fluorescence Microscopy," Optics Letters 19, 780-782 (1994).
[47] G. Moneron and S. W. Hell, "Two-photon excitation STED microscopy," Opt Express 17, 14567-14573 (2009).
[48] V. Westphal and S. W. Hell, "Nanoscale resolution in the focal plane of an optical microscope," Phys Rev Lett 94, 143903-143906(2005).
[49] J. A. J. Fitzpatrick, Q. Yan, J. J. Sieber, M. Dyba, U. Schwarz, C. Szent-Gyorgyi, C. A. Woolford, P. B. Berget, A. S. Waggoner, and M. P. Bruchez, "STED Nanoscopy in Living Cells Using Fluorogen Activating Proteins," Bioconjugate Chem 20, 1843-1847 (2009).
[50] E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, "Imaging Intracellular Fluorescent Proteins at Nanometer Resolution," Science 313, 1642-1645 (2006).
[51] S. T. Hess, T. P. K. Girirajan, and M. D. Mason, "Ultra-high resolution imaging by fluorescence photoactivation localization microscopy," Biophys J 91, 4258-4272 (2006).
[52] M. J. Rust, M. Bates, and X. W. Zhuang, "Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM)," Nat Methods 3, 793-795 (2006).
[53] B. Huang, W. Wang, M. Bates, and X. Zhuang, "Three-Dimensional Super-Resolution Imaging by Stochastic Optical Reconstruction Microscopy," Science 319, 810-813 (2008).
[54] M. Heilemann, P. Dedecker, J. Hofkens, and M. Sauer, "Photoswitches: Key molecules for subdiffraction-resolution fluorescence imaging and molecular quantification," Laser Photonics Rev 3, 180-202 (2009).
[55] M. G. L. Gustafsson, "Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy," J Microsc-Oxford 198, 82-87 (2000).
[56] M. G. L. Gustafsson, "Extended resolution fluorescence microscopy," Curr Opin Struc Biol 9, 627-634 (1999).
[57] R. Heintzmann, T. M. Jovin, and C. Cremer, "Saturated patterned excitation microscopy concept for optical resolution improvement," J. Opt. Soc. Am. A 19, 1599-1609 (2002).
[58] R. Heintzmann, "Saturated patterned excitation microscopy with two-dimensional excitation patterns," Micron 34, 283-291 (2003).
[59] M. G. L. Gustafsson, L. Shao, P. M. Carlton, C. J. R. Wang, I. N. Golubovskaya, W. Z. Cande, D. A. Agard, and J. W. Sedat, "Three-dimensional resolution doubling in wide-field fluorescence microscopy by structured illumination," Biophys J 94, 4957-4970 (2008).
[60] R. Ayuk, H. Giovannini, A. Jost, E. Mudry, J. Girard, T. Mangeat, N. Sandeau, R. Heintzmann, K. Wicker, K. Belkebir, and A. Sentenac, "Structured illumination fluorescence microscopy with distorted excitations using a filtered blind-SIMalgorithm," Optics Letters 38, 4723-4726 (2013).
[61] J. Lu, W. Min, J. A. Conchello, X. S. Xie, and J. W. Lichtman, "Super-Resolution Laser Scanning Microscopy through Spatiotemporal Modulation," Nano Lett 9, 3883-3889 (2009).
[62] K. Isobe, T. Takeda, K. Mochizuki, Q. Song, A. Suda, F. Kannari, H. Kawano, A. Kumagai, A. Miyawaki, and K. Midorikawa, "Enhancement of lateral resolution and optical sectioning capability of two-photon fluorescence microscopy by combining temporal-focusing with structured illumination," Biomed. Opt. Express 4, 2396-2410 (2013).
[63] P. W. Winter, P. Chandris, R. S. Fischer, Y. Wu, C. M. Waterman, and H. Shroff, "Incoherent structured illumination improves optical sectioning and contrast in multiphoton super-resolution microscopy," Opt Express 23, 5327-5334 (2015).
[64] A. Punge, S. O. Rizzoli, R. Jahn, J. D. Wildanger, L. Meyer, A. Schonle, L. Kastrup, and S. W. Hell, "3D reconstruction of high-resolution STED microscope images," Microsc Res Techniq 71, 644-650 (2008).
[65] H. S. Zahra and H. T. Oweis, "Application of high-pass filtering techniques on gravity and magnetic data of the eastern Qattara Depression area, Western Desert, Egypt," NRIAG Journal of Astronomy and Geophysics.
[66] P. Yeh, Optical Waves in Layered Media (John Wiley & Sons., Inc, Canada,1988).
[67] C.-H. Yeh and S.-Y. Chen, "Resolution enhancement of two-photon microscopy via intensity-modulated laser scanning structured illumination," Appl Optics 54, 2309-2317 (2015).
[68] R. C. Gonzalez and R. E. Woods, Digital Image Processing (Addison-Wesley Longman Publishing Co., Inc., New Jersey, 2001).
[69] S. A. Shroff, J. R. Fienup, and D. R. Williams, "Phase-shift estimation in sinusoidally illuminated images for lateral superresolution," Journal of the Optical Society of America A 26, 413-424 (2009).
[70] K. Wicker, O. Mandula, G. Best, R. Fiolka, and R. Heintzmann, "Phase optimisation for structured illumination microscopy," Opt Express 21, 2032-2049 (2013).
[71] M. Muller, V. Monkemoller, S. Hennig, W. Hubner, and T. Huser, "Open-source image reconstruction of super-resolution structured illumination microscopy data in ImageJ," Nat Commun 7(2016).
[72] A. Lal, C. Shan, and P. Xi, "Structured Illumination Microscopy Image Reconstruction Algorithm," Ieee J Sel Top Quant 22, 1-14 (2016).
[73] C. H. Yeh, C. Z. Tan, and S. Y. Chen, “Improved Resolution of Second Harmonic Generation Microscopy in Deep Tissue via Structured Scanning Pattern,” manuscript in progress

指導教授

陳思妤(Szu-Yu Chen)

審核日期

2016-7-28

推文