目前多光子點掃描式(multiphoton-induced point-scanning)加工方法雖然可以達到任意形狀之三維(three-dimensional，3D)非熱致效應的加工效果，但是因為其採用點對點掃描的加工方式而使得其加工速度較為緩慢，而為了克服這個限制，本論文使用了同時空間與時間聚焦(spatial and temporal focusing)的廣視域(wide-field)高通量(high-throughput)之多光子激發技術來完成生物組織加工。
在本論文中使用鈦藍寶石雷射再生放大器(Ti:sapphire regenerative amplifier)作為廣視域系統的激發光源，再搭配繞射光柵(diffraction grating)、高數值孔徑(numerical aperture)物鏡、準直透鏡(collimated lens)、光柵脈衝壓縮器(grating pulse compressor)、干涉式自相關儀(interference autocorrelator)、電荷耦合元件(charge coupled device)、三軸電控移動平台(three-axis motorized stage)，使其具有廣視域激發與獲取影像的能力。在本系統中，激發光直徑可達到80 μm，且單發雷射脈衝之能量密度最高可達到4.5 J/cm2。而藉由多光子誘發蝕除(multiphoton-induced ablation)技術，目前初步完成了任意二維圖形之微加工而加工時間只僅僅需要幾秒鐘，與傳統多光子單點掃描加工方式相互比較，其加工速度提升了約為100倍。在本論文中也試圖對生物組織如雞腱，進行高速的多光子誘發蝕除，藉以呈現系統的高通量特性，此大範圍之高速多光子誘發蝕除技術可以利用在全光組織學(all-optical histology)上，藉由迭代的高通量之蝕除與影像掃描來獲得全組織的內部膠原蛋白的二倍頻(second harmonic generation，SHG)影像影像。

Conventional multiphoton-induced point-scanning methods have been successfully utilized to process arbitrary three-dimensional (3D) patterns without thermal damage in adjacent part; however, one of the limits in this approach is its low throughout due to point-by-point processing. To break this limit, a high- throughput and wide-field ablation system based on spatiotemporal focusing has been developed to rapidly remove bio-tissues.
In this thesis, we have developed high-throughput and wide-field ablation system based on spatiotemporal focusing. The key components include a Ti: sapphire regenerative amplifier laser as the excitation source, a diffraction grating, a high numerical aperture objective, a collimated lens, a grating pulse compressor, an interference autocorrelator, a three-axis motorized stage, and a charge coupled device (CCD) to achieved wide-field excitation and imaging. The excited beam’s diameter on the sample is around 80 μm and laser energy density can be up to 4.5 J/cm2 per pulse. In the two-dimensional patternized ablation, the ablation time for machining the micro-pattern is only a few seconds. Compared with the conventional multiphoton-induced point-scanning method, this approach increases the ablating speed more than two orders. In order to perform the high-throughput characteristics of this system, we also tried high-speed and large-area ablation of biological tissues such as chicken tendon; this technique can be used in the all-optical histology, by high-throughput iteratively ablation and imaging to obtain second-harmonic generation image from the collagen within the biological specimen.

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