小動物常用來做為研究各種疾病的實驗室模型，小動物影像能縮短相關研究及藥物開發時程，提升研究的品質，然而其較臨床醫學影像需要更高的解析度及影像對比。此外，近來對於開發能顯示觀察病理過程中微細血管脈絡生長及重建的相關技術需求也有愈來愈高的趨勢。在本研究中，我們建立了一套非侵入式高頻暗場共軛焦光聲顯微影像系統，其透過光聲效應來偵測被組織吸收的光子，基於組織中血液本身對於雷射光的高吸收對比及透過高頻超音波的偵測方式，結合了光學和聲學兩者的優點以提供高對比及高空間解析度，可用來進行小動物微細血管脈絡造影。此系統可工作於25 MHz及50 MHz兩組工作頻率，可提供B-scan與C-scan兩種掃瞄模式。當工作頻率為25 MHz時，此系統可提供68 um軸向解析度、171 um橫向解析度及至少6 mm的穿透深度；當工作在50 MHz的頻率時，可提供高達36 um的軸向解析度、65 um橫向解析度及至少3 mm的穿透深度。此系統亦可進行高頻超音波掃瞄，以提供資訊互補之傳統超音波B-mode結構性影像。此系統並經活體小動物實驗驗證，確能提供高解析度及高對比小鼠皮下微細血管脈絡及腫瘤血管分布影像。本研究並嘗試開發此光聲顯微影像系統於生醫研究上的新應用，將其用於小鼠損傷的阿基里斯腱內微細血管增生的監控上，以及搭配可調光吸收波長之金奈米桿粒子作為光聲造影對比劑，利用金奈米桿粒子對近紅光的強吸收及其因奈米尺寸大小而易由血腦屏障開啟區域滲入腦組織的特性，更進一步地將此系統應用監控小動物模型之聚焦式超音波誘發血腦屏障開啟過程，驗證其可行性。未來將持續改善系統效能與活體實驗的流程，期望未來能提供運動醫學及血腦屏障開啟等相關診療及研究一個新的、有用的影像工具。

Small animals are the preferred laboratory models for studying various diseases. Small animal imaging provides the opportunity to evaluate pathologic progression in a much-compressed time frame while it requires much higher resolution and image contrast than clinical medical imaging. Moreover, there is a significant need to develop technologies for visualizing micro-vascular growth and remodeling in pathogenic process. In this study, we developed a non-invasive, high frequency dark-field confocal photoacoustic microscope (PAM) system. PAM, a hybrid biophotonic imaging technique detecting absorbed photons ultrasonically through the photoacoustic effect, has the potential to address these needs mentioned above. With strong endogenous blood optical absorption contrast and high frequency ultrasound detection, PAM can provide high contrast and high spatial resolution for micro-vasculature imaging of small animals. Currently, we have developed 25-MHz and 50-MHz dark-field confocal PAM systems. Our PAM system provides two imaging modes – B-scan (2D imaging) and C-scan (3D imaging). When using a 25-MHz ultrasonic transducer, its maximum achievable resolution is 68 um in axial and 171 um in lateral, and the penetration depth can reach at least 6 mm. With a 50-MHz transducer, higher resolution can be achieved – 36 um in axial and 65 um in lateral while compromising with penetration depth – at least 3 mm. The performance of our PAM is further verified with in vivo small animal experiments – high resolution and high contrast subcutaneous and tumor micro-vasculature imaging of mice can be obtained. We also tried to explore new biomedical applications of our PAM, applying it to image micro-vascular changes in injured Achilles tendons of mice. In addition, by taking advantage of the strong near-infrared absorption of gold nanorods (AuNRs) and their extravasation tendency from blood-brain barrier (BBB) opening foci due to their nano-scale sizes, we employed our PAM system along with AuNRs as photoacoustic contrast agents to monitor focused-ultrasound induced BBB opening in small animal models in vivo, verifying the feasibility. Future work will focus on optimization of our PAM system and in vivo small animal experimental procedures for the tendon and BBB-opening imaging. Our preliminary studies show promising of photoacoustic imaging as a useful aid of sports medicine and BBB opening monitoring.

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