Abstract
Two-photon excitation laser scanning fluorescence microscopy (TPLSM) is a powerful tool to visualize intravital microstructures. This is because of its superior penetration depth and less-invasiveness in specimens owing to its near-infrared excitation laser wavelength compared with the single-photon excitation. In this presentation, our studies to improve spatial and temporal resolution of TPLSM by utilizing some of novel optical technologies are introduced. To improve the spatial resolution, we applied stimulated emission depletion (STED) technologies utilizing electrically controllable components, transimissive liquid crystal devises and laser diode-based light sources [Opt. Express 2014; Biomed Opt. Express 2018], resulting in 5-times hihger spatial resolution than conventional [Biomed Opt. Express 2019]. In addition, we recently adopted other technologies such as adaptive optics [PLOS One 2020; ACS Omega 2021] or nano-sheet based observation methods [iSci. 2020; Star Protocol's. 2021] for sharper intravital imaging. For higher temporal resolution, a confocal spinning disk scanning unit and high-peak-power laser light sources were utilized for TPLSM imaging [Anal. Sci. 2015; Front. Physical. 2019; BBRC 2020]. The developed system achieved large field of view, fine spatial resolution and penetration depth, and has been used for several biological applications.
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