Chen Dihan, Ren Mindan, Zhang Dapeng, Chen Jialong, Gu Songyun, Chen Shih-Chi
Opt Express. 2020 Sep 28;28(20):30187-30198. doi: 10.1364/OE.404652.
We present the modular design and characterization of a multi-modality video-rate two-photon excitation (TPE) microscope based on integrating a digital micromirror device (DMD), which functions as an ultrafast beam shaper and random-access scanner, with a pair of galvanometric scanners. The TPE microscope system realizes a suite of new imaging functionalities, including (1) multi-layer imaging with 3D programmable imaging planes, (2) DMD-based wavefront correction, and (3) multi-focus optical stimulation (up to 22.7 kHz) with simultaneous TPE imaging, all in real-time. We also report the detailed optomechanical design and software development that achieves high level system automation. To verify the performance of different microscope functions, we have devised and performed imaging experiments on Drosophila brain, mouse kidney and human stem cells. The results not only show improved imaging resolution and depths via the DMD-based adaptive optics, but also demonstrate fast multi-focus stimulation for the first time. With the new imaging capabilities, e.g., tools for optogenetics, the multi-modality TPE microscope may play a critical role in the applications pertinent to neuroscience and biophotonics.
我们展示了一种基于数字微镜器件（DMD）与一对振镜集成的多模态视频速率双光子激发（TPE）显微镜的模块化设计与特性。DMD作为超快光束整形器和随机访问扫描仪，与一对振镜集成在一起。该TPE显微镜系统实现了一系列新的成像功能，包括：（1）具有3D可编程成像平面的多层成像；（2）基于DMD的波前校正；（3）高达22.7kHz的多焦点光学刺激与同步TPE成像，所有这些均为实时操作。我们还报告了实现高级系统自动化的详细光机械设计和软件开发过程。为了验证不同显微镜功能的性能，我们设计并在果蝇大脑、小鼠肾脏和人类干细胞上进行了成像实验。结果不仅通过基于DMD的自适应光学技术提高了成像分辨率和深度，还首次展示了快速多焦点刺激。凭借诸如光遗传学工具等新的成像能力，多模态TPE显微镜可能在神经科学和生物光子学相关应用中发挥关键作用。
