Journal
PLOS ONE
Volume 10, Issue 1, Pages -Publisher
PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0116280
Keywords
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Categories
Funding
- JSPS KAKENHI grants of the Ministry of Education, Culture, Sports, Science and Technology, Japan [26242082, 22113005]
- Nano-Macro Materials, Devices and System Research Alliance
- Network Joint Research Center for Materials and Devices
- [26-1533]
- Grants-in-Aid for Scientific Research [22113005, 25560411, 26242082, 26650107, 14J01533] Funding Source: KAKEN
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Elucidation of neural circuit functions requires visualization of the fine structure of neurons in the inner regions of thick brain specimens. However, the tissue penetration depth of laser scanning microscopy is limited by light scattering and/or absorption by the tissue. Recently, several optical clearing reagents have been proposed for visualization in fixed specimens. However, they require complicated protocols or long treatment times. Here we report the effects of 2,2'-thiodiethanol (TDE) solutions as an optical clearing reagent for fixed mouse brains expressing a yellow fluorescent protein. Immersion of fixed brains in TDE solutions rapidly (within 30 min in the case of 400-mu m-thick fixed brain slices) increased their transparency and enhanced the penetration depth in both confocal and two-photon microscopy. In addition, we succeeded in visualizing dendritic spines along single dendrites at deep positions in fixed thick brain slices. These results suggest that our proposed protocol using TDE solution is a rapid and useful method for optical clearing of fixed specimens expressing fluorescent proteins.
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