由於多光子激發螢光顯微鏡(multi-photon excited fluorescence microscopy，MPEFM)系統的深度成像能力及高空間分辨率特性，目前已成為生物組織研究中不可或缺的重要工具；而為進一步提升螢光影像之解析度與對比度，本論文將其與適應性光學(adaptive optics，AO)概念相結合，預先補償系統及樣品上的像差。
本論文首先建立一套AO系統，利用15項Zernike多項式(Zernike polynomial)作為數學模型，並分別藉由自製Shack-Hartmann波前感測器(Shack-Hartmann wavefront sensor，SHWS)與可調變式聚焦鏡(deformable mirror，DM)作為波前感測器及波前修正元件，接著結合CameraLink高速傳輸介面及現場可程式化邏輯閘陣列(field programmable gate array，FPGA)的即時運算能力，使系統達到200 Hz之修正像差效果；此外，由於是利用平行化PI控制器(parallel PI controller)，藉由Zernike多項式各像差間相互正交的特性對DM產生的面形進行控制，因此能夠單獨調控各像差，使系統於修正干擾同時控制聚焦點於光軸上的位置，達成快速遙控調焦(remote focusing，RF)之目標。
最後本論文將AO概念與MPEFM系統整合，同樣以DM作為波前修正元件，藉由分析螢光影像強度，逐一調整DM產生出的各項像差，找尋出修正干擾的最佳Zernike係數組合，達成非感測試AO (sensorless AO)。

The multi-photon excitation fluorescence microscopy (MPEFM) system is an advanced and powerful imaging technology widely employed in the fields of life sciences and biomedical research. By utilizing the non-linear optical process of multi-photon excitation, it offers high-resolution imaging, deep tissue penetration, and three-dimensional reconstruction capabilities. In this study, we incorporated the concept of adaptive optics (AO) into the MPEFM system to achieve aberration correction and remote focusing (RF). This was accomplished by leveraging the high response rate of a deformable mirror (DM) and the orthogonal properties of Zernike polynomials, which represent various aberrations. Before performing aberration correction, a three-step identification process was conducted to ensure precise compensation of individual aberrations by the DM. The results demonstrated that this process yielded a signal-to-noise (SNR) ratio exceeding 20.3 dB for all aberrations, without regarding the x-tilt and y-tilt terms. In the final experiment, our adaptive optics system exhibited a reduction in wavefront variance by approximately 475-fold and enabled a remote focusing range of 128 µm. Lastly, the integration of the AO system into the MPEFM system led to a more than three-fold increase in fluorescence image intensity.

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