Nanoscale mapping of bio-molecular building blocks of brain


Zhuyu Peng and Jinyoung Kang

Zhuyu Peng and Jinyoung Kang

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For understanding the brain structure, it is necessary to decipher the nanoscale organization of the biomolecular building blocks of the brain in 3D. However, the resolution of optical microscopes is limited by the diffraction of light while conventional super-resolution technologies require specialized expensive equipment and have challenges in scaling to 3D [K. R. Porter et. al., J. Exp. Med., 81, 233–246 (1945)] [E. Betzig, E. et al., Science, 313, 1642–1645 (2006)] [M. J. Rust et. al., Nat Methods, 3, 793–795 (2006)]. Recently, it has been shown that it is possible to break the diffraction limited resolution of optical microscopes by physically expanding the biological samples using electrostatic forces in a hydrogel: a methodology named expansion microscopy (ExM) [F. Chen et. al., Science, 347 (6221), 543–548 (2015)]. We have developed the technology to achieve the highest expansion factor, reported till date (100-fold linear expansion), of tissue-polymer hybrids [D. Sarkar et. al., Society for Neuroscience (2016)] . Such high physical expansion factors allow imaging of biological specimens at sub-10 nm resolution (i.e., 300 nm (diffraction limit) / 100 (expansion factor)), using conventional diffraction limited microscopes. This technology, which we termed Expansion Revealing (ExR), utilizes both electrostatic and mechanical forces to achieve extremely high expansion factors and is fundamentally different from expansion microscopy (ExM), or iterated-ExM, which involves solely the electrostatic repulsive forces in the polymer for expansion [D. Sarkar et. al., Nature Biomedical Engg., 1-17 (2022)]. Moreover, ExR is much simpler to implement, provides better yield and retention of biomolecules, allowing post processing. ExR enables precise mapping of the biomolecular building blocks of cells (proteins, transcriptomes (RNA), DNA) as well as the cellular interconnections that form large scale, 3D circuits, using hardware and reagents easily available in research laboratories. Thus, it is highly advantageous compared to conventional super-resolution imaging techniques, which are difficult to scale to 3D thick tissues and require forbiddingly expensive hardware and expert handling.

We are now working on applying this technology for mapping the biomolecular building blocks of brain and provide in-depth insights into the pathology mechanisms that could lead to the discovery of new targets for treating neurological diseases.