Super-Resolution Imaging in Collagen-Abundant Thick Tissues

Authors:

Ya-Han Chuang, Yueh-Feng Wu, Ya-Hui Lin, Yin-Hsu Chen, Yu-Xian Zhou, Shao-Chun Hsu, Hsin-Mei Lee, Ann-Shyn Chiang, Yunching Chen, Shiang-Jiuun Chen, Sung-Jan Lin, Li-An Chu

Keywords:

Expansion microscopy; ExM; collagen ExM, ColExM; physical expansion; super-resolution microscopy

Abstract:

Expansion microscopy (ExM) has gained increasing popularity for 3D ultrastructural imaging of cultured cells and tissue slices at nanoscale resolution using conventional microscopes via physical expansion of biological tissues. However, its application to collagen-abundant thick tissues is still challenging. Herein, a new method, collagen ExM (ColExM), optimized for expanding tissues containing more than 70% collagen, is demonstrated. ColExM succeeds in 4.5-fold linear expansion with minimal structural distortion of corneal and skin tissues. It is compatible with immunostaining, allowing super-resolution visualization of 3D neural structures innervating hair follicles, corneas, and pancreatic tumors with high stromal collagen content. The method succeeds in identifying individual mitochondria and previously unrecognized dendritic spinelike structures of corneal nerves. It also enables fine mapping of structural rearrangement of tight junctions and actin cytoskeletons. Therefore, ColExM can facilitate the exploration of 3D nanoscale structures in collagen-rich tissues.

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