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Low-invasive 5D visualization of mitotic progression by two-photon excitation spinning-disk confocal microscopy
| Title: | Low-invasive 5D visualization of mitotic progression by two-photon excitation spinning-disk confocal microscopy |
| Authors: | Kamada, Takafumi Browse this author | | Otomo, Kohei Browse this author →KAKEN DB | | Murata, Takashi Browse this author →KAKEN DB | | Nakata, Kaito Browse this author | | Hiruma, Shota Browse this author | | Uehara, Ryota Browse this author →KAKEN DB | | Hasebe, Mitsuyasu Browse this author | | Nemoto, Tomomi Browse this author →KAKEN DB |
| Issue Date: | 17-Jan-2022 |
| Publisher: | Nature Portfolio |
| Journal Title: | Scientific reports |
| Volume: | 12 |
| Issue: | 1 |
| Start Page: | 809 |
| Publisher DOI: | 10.1038/s41598-021-04543-7 |
| Abstract: | Non-linear microscopy, such as multi-photon excitation microscopy, offers spatial localities of excitations, thereby achieving 3D cross-sectional imaging with low phototoxicity even in thick biological specimens. We had developed a multi-point scanning two-photon excitation microscopy system using a spinning-disk confocal scanning unit. However, its severe color cross-talk has precluded multi-color simultaneous imaging. Therefore, in this study, we introduced a mechanical switching system to select either of two NIR laser light pulses and an image-splitting detection system for 3- or 4-color imaging. As a proof of concept, we performed multi-color fluorescent imaging of actively dividing human HeLa cells and tobacco BY-2 cells. We found that the proposed microscopy system enabled time-lapse multi-color 3D imaging of cell divisions while avoiding photodamage. Moreover, the application of a linear unmixing method to the 5D dataset enabled the precise separation of individual intracellular components in multi-color images. We thus demonstrated the versatility of our new microscopy system in capturing the dynamic processes of cellular components that could have multitudes of application. |
| Type: | article |
| URI: | http://hdl.handle.net/2115/84188 |
| Appears in Collections: | 電子科学研究所 (Research Institute for Electronic Science) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
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