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RESEARCH ARTICLE
Year : 2022  |  Volume : 17  |  Issue : 6  |  Page : 1348-1356

Optical tissue clearing enables rapid, precise and comprehensive assessment of three-dimensional morphology in experimental nerve regeneration research


1 SickKids Research Institute, Neuroscience and Mental Health Program, Toronto, ON, Canada
2 SickKids Research Institute, Neuroscience and Mental Health Program; Division of Plastic and Reconstructive Surgery, the Hospital for Sick Children, Toronto, ON, Canada
3 SickKids Research Institute, Neuroscience and Mental Health Program; Institute of Biomaterials and Biomedical Engineering, University of Toronto; Division of Plastic and Reconstructive Surgery, the Hospital for Sick Children, Toronto, ON, Canada; Indiana University School of Medicine, Indianapolis, IN, USA

Correspondence Address:
Simeon C Daeschler
SickKids Research Institute, Neuroscience and Mental Health Program, Toronto, ON
Canada
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Source of Support: This work was supported by the German Research Foundation (DA 2255/1-1, to SCD), Conflict of Interest: None


DOI: 10.4103/1673-5374.329473

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Morphological analyses are key outcome assessments for nerve regeneration studies but are historically limited to tissue sections. Novel optical tissue clearing techniques enabling three-dimensional imaging of entire organs at a subcellular resolution have revolutionized morphological studies of the brain. To extend their applicability to experimental nerve repair studies we adapted these techniques to nerves and their motor and sensory targets in rats. The solvent-based protocols rendered harvested peripheral nerves and their target organs transparent within 24 hours while preserving tissue architecture and fluorescence. The optical clearing was compatible with conventional laboratory techniques, including retrograde labeling studies, and computational image segmentation, providing fast and precise cell quantitation. Further, optically cleared organs enabled three-dimensional morphometry at an unprecedented scale including dermatome-wide innervation studies, tracing of intramuscular nerve branches or mapping of neurovascular networks. Given their wide-ranging applicability, rapid processing times, and low costs, tissue clearing techniques are likely to be a key technology for next-generation nerve repair studies. All procedures were approved by the Hospital for Sick Children’s Laboratory Animal Services Committee (49871/9) on November 9, 2019.


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