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Long Preservation of AAV-Transduced Fluorescence by a Modified Organic Solvent-Based Clearing Method
Tao Lu1, Munehisa Shinozaki1, Narihito Nagoshi2
1Department of Physiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan.
International Journal of Molecular Sciences
|September 9, 2022
Summary
This study introduces tDISCO, a modified tissue clearing method. It preserves fluorescence and rapidly clears tissues for enhanced 3D imaging of neural projections.
Area of Science:
- Neuroscience
- Biotechnology
- Microscopy
Background:
- Tissue clearing enables 3D imaging of whole organs, crucial for studying the central nervous system.
- Existing methods like 3DISCO face limitations due to fluorescence quenching and peroxide removal.
Purpose of the Study:
- To develop a modified tissue clearing protocol (tDISCO) that overcomes limitations of existing methods.
- To enable long-term preservation of endogenous fluorescence and simplify the clearing process.
Main Methods:
- A modified tissue clearing protocol, termed tDISCO, was developed.
- The protocol was tested for its ability to preserve fluorescence and achieve transparency.
- tDISCO was applied to various tissues, including brain, other organs, and bone.
Main Results:
- The tDISCO protocol preserves AAV-transduced EGFP fluorescence for extended periods (months).
- Achieved high transparency with a fast and simple clearing process.
- Successfully visualized long projection neurons and axons in 3D across the brain and spinal cord.
Conclusions:
- tDISCO offers a practical and efficient solution for 3D visualization of neural structures.
- This method enhances the study of neuronal projections throughout the central nervous system.
- tDISCO is applicable to diverse tissues, including challenging samples like bone.
