Three-photon microscopy (3PM) excited at both the 1300 and 1700 nm window are established deep-brain imaging technologies. Beyond 1700 nm lies the last unexplored optical window potential for deep-brain 3PM: the 2200 nm window. However, there has been no experimental demonstration of brain 3PM at this window, and hence no quantitative comparison with other excitation windows. Here we demonstrate brain 3PM excited at 2200 nm in vivo, by developing an energetic femtosecond laser and a laser scanning microscope with high transmittance and minimum pulse broadening. We also perform 3PM excited at 1700 nm for direct comparison. Our results show that the effective attenuation length, a measure of the maximum achievable imaging depth, at 2200 nm is smaller than that at 1700. Besides, the spatial resolution of 2200 nm 3PM is lower than that of 1700 nm 3PM. We thus fill this last missing gap in deep-brain MPM, in terms of excitation wavelength selection.

中文翻译：

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小鼠大脑在 2200 nm 窗口激发的体内三光子显微镜

在 1300 和 1700 nm 窗口激发的三光子显微镜 (3PM) 是成熟的脑深部成像技术。超过 1700 nm 是深脑 3PM 的最后一个未开发的光学窗口潜力：2200 nm 窗口。然而，在这个窗口没有大脑 3PM 的实验证明，因此没有与其他激发窗口的定量比较。在这里，我们通过开发高能飞秒激光器和具有高透射率和最小脉冲展宽的激光扫描显微镜，展示了在 2200 nm 体内激发的大脑 3PM。我们还进行了 1700 nm 激发的 3PM 以进行直接比较。我们的结果表明，2200 nm 处的有效衰减长度（衡量最大可实现成像深度）小于 1700 nm。此外，2200 nm 3PM 的空间分辨率低于 1700 nm 3PM。