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An Improved SHANEL Procedure for Clearing and Staining Brain Tissue from Multiple Species
Fu Zeng1,2, Lian Huang1,2, Ding Han3
1State Key Laboratory of Common Mechanism Research for Major Diseases, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine Peking Union Medical College, Beijing 100005, China.
International Journal of Molecular Sciences
|May 7, 2025
Summary
We developed iSHANEL, an improved tissue clearing method for high-resolution 3D brain imaging. This technique enhances transparency and visualization in diverse species, overcoming limitations of previous methods.
Area of Science:
- Neuroscience
- Biotechnology
- Microscopy
Background:
- Tissue clearing enables 3D imaging of whole organs, but faces challenges with thick human brain samples.
- Limitations include inefficient clearance, long processing times, and autofluorescence from pigments and blood in formalin-fixed tissues.
Purpose of the Study:
- To develop an improved tissue clearing procedure, iSHANEL, for enhanced 3D brain imaging across multiple species.
- To address limitations of existing methods, including lipid removal and autofluorescence reduction.
Main Methods:
- Developed and applied the iSHANEL (improved SHANEL) procedure to brain tissue from various species, including non-human primates.
- Evaluated lipid removal, particularly sphingolipids, and assessed tissue transparency and protein immunogenicity.
- Investigated methods to reduce blood vessel autofluorescence.
Main Results:
- iSHANEL significantly enhanced tissue transparency and effectively removed lipids across species.
- Improved visualization of brain vasculature with reduced light scattering and background noise.
- Demonstrated successful application to large pig and cynomolgus monkey brains, preserving protein integrity (IBA-1 staining).
Conclusions:
- iSHANEL offers an improved method for high-resolution 3D imaging of brain tissue.
- The procedure is compatible with diverse species and addresses key limitations of prior tissue clearing techniques.
- iSHANEL facilitates detailed visualization of brain structures and vasculature.
