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Published on: October 8, 2013
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A rapid optical clearing protocol using 2,2'-thiodiethanol for microscopic observation of fixed mouse brain
Yuka Aoyagi1, Ryosuke Kawakami2, Hisayuki Osanai1
1Research Institute for Electronic Science, Hokkaido University, Sapporo, Hokkaido, Japan; Graduate School of Information Science and Technology, Hokkaido University, Sapporo, Hokkaido, Japan.
Plos One
|January 31, 2015
Summary
Optical clearing of thick brain slices is crucial for visualizing neural circuits. Researchers found that 2,2′-thiodiethanol (TDE) solutions rapidly clear fixed brain tissue, enhancing microscopy penetration depth for detailed neuronal structure imaging.
Area of Science:
- Neuroscience
- Microscopy
- Biochemistry
Background:
- Visualizing neural circuits requires imaging fine neuronal structures in thick brain specimens.
- Laser scanning microscopy faces limitations in tissue penetration depth due to light scattering and absorption.
- Existing optical clearing reagents often involve complex protocols or lengthy treatment times.
Purpose of the Study:
- To evaluate 2,2′-thiodiethanol (TDE) solutions as a rapid optical clearing reagent for fixed mouse brain specimens.
- To assess the effectiveness of TDE in enhancing transparency and microscopy penetration depth.
- To demonstrate the utility of TDE for visualizing deep neuronal structures.
Main Methods:
- Fixed mouse brains expressing yellow fluorescent protein were immersed in TDE solutions.
- Transparency and penetration depth were evaluated using confocal and two-photon microscopy.
- The ability to visualize fine structures like dendritic spines at depth was assessed.
Main Results:
- TDE solutions rapidly increased the transparency of fixed brain slices within 30 minutes.
- Enhanced transparency led to improved light penetration depth in microscopy.
- Successful visualization of dendritic spines along dendrites in deep tissue sections was achieved.
Conclusions:
- TDE solutions offer a fast and effective method for optical clearing of fixed fluorescent protein-expressing specimens.
- This protocol significantly improves deep tissue imaging capabilities in microscopy.
- The TDE method facilitates detailed structural analysis of neural circuits in thick brain samples.
