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標題: | 雙光子聲光顯微術的實現與架設 Realization and Implementation of Two-photon Acoustic Microscopy |
作者: | Szu-Yu Lee 李思宇 |
指導教授: | 孫啟光(Chi-Kuang Sun) |
關鍵字: | 聲光顯微術,鈦藍寶石雷射,雙光子吸收,光學調變,次微米解析度,活體影像,黑色素分布, Photoacoustic microscopy,Ti:sapphire laser,two-photon absorption,optical modulation,sub-micron resolution,in-vivo imaging,melanin distribution, |
出版年 : | 2015 |
學位: | 碩士 |
摘要: | 聲光顯微鏡是一種以光學吸收為對比來源的新穎生物醫療用顯微術,乃針對非輻散的信號進行偵測的成像系統。該顯微術可以支援不同尺度的視野觀察,同時提供樣本的多種生物資訊。在針對細胞到胞器尺度的觀察應用上,高解析度的聲光顯微術有其不可取代的價值,而在這種微米到次微米的視野大小下,一般的高解析聲光顯微術並不能同時在徑向和軸向上達到微米等級的解析度,此外,其成像速度相當緩慢,這兩大因素造成高解析聲光顯微術在許多生物或臨床應用上的限制。為了解決目前高解析聲光顯微術所遇到的瓶頸,在此論文中,我們提出一種使用不同於一般聲光顯微術單光子吸收機制的作法。我們研發了一種以雙光子吸收為機制的雷射掃描聲光顯微鏡,可以應用於活體實驗中即時的影像觀察。此顯微術以鈦藍寶石飛秒雷射作為光源,激發生物組織中的雙光子吸收致聲光信號,由於此非線性的光學物理機制,可以同時滿足次微米等級的徑向解析度和微米等級的軸向解析度,達到超高的三維空間解析度。
本論文從頭開始討論如何善用一套光學調變系統,實現此雙光子聲光顯微術,同時探討有效的雙光子聲光對比劑,並最佳化此聲光顯微系統。接下來,針對此顯微術的徑向和軸向空間解析度分別進行量測。最後,為了達到即時成像速度,我們把聲光成像系統和掃描式顯微鏡做整合,完成這套高速掃描式雙光子聲光顯微鏡,並成功示範此顯微術應用於活體老鼠皮膚的黑色素觀察。從該活體實驗中,我們證明了此技術能夠克服當前高解析聲光顯微術的瓶頸,並可以支援許多生物和臨床上的應用。 As a new branch of optical-absorption based microscopy with detection of non-radiative emission, photoacoustic microscopy provides manifold bio-information with scalable field of view inside biological specimens. For cell or organelle micro-level imaging, high resolving capability is necessary. However, among common photoacoustic microscopy, high lateral and axial resolution is achieved separately by different engineering methods. Furthermore, imaging rate is relatively slow compared to other in-vivo imaging modalities. To simultaneously obtain high spatial resolution in three dimensions, in this thesis, I report an in-vivo photoacoustic microscopy by a distinctive physical mechanism of contrast − two-photon absorption. The light source is a Ti:sapphire laser with wavelength at 800~860 nm. Due to the nonlinear absorption property, intrinsic photoacoustic signals confinement ensures the high lateral and axial resolution, all together. Starting from scratch, I developed and fully studied a loss modulation technique to realize this two-photon acoustic microscopy, investigated potential contrast agents, characterized spatial resolution, enhanced imaging speed by integration with a commercial scanning microscope, and demonstrated biological applications. 0.52 μm lateral and 1.94 μm axial resolution is achieved with high imaging rate, 0.38fps (512×512 pixels). Finally, I utilized this two-photon acoustic microscopy to demonstrate label-free imaging of non-fluorescent melanin distribution within a black mouse ear both ex-vivo and in-vivo. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/54664 |
全文授權: | 有償授權 |
顯示於系所單位: | 光電工程學研究所 |
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