Obtaining high-resolution information from a complex system, while maintaining the global perspective needed to understand system function, represents a key challenge in biology. Here we address this challenge with a method (termed CLARITY) for the transformation of intact tissue into a nanoporous hydrogel-hybridized form (crosslinked to a three-dimensional network of hydrophilic polymers) that is fully assembled but optically transparent and macromolecule-permeable. Using mouse brains, we show intact-tissue imaging of long-range projections, local circuit wiring, cellular relationships, subcellular structures, protein complexes, nucleic acids and neurotransmitters. CLARITY also enables intact-tissue in situ hybridization, immunohistochemistry with multiple rounds of staining and de-staining in non-sectioned tissue, and antibody labelling throughout the intact adult mouse brain. Finally, we show that CLARITY enables fine structural analysis of clinical samples, including non-sectioned human tissue from a neuropsychiatric-disease setting, establishing a path for the transmutation of human tissue into a stable, intact and accessible form suitable for probing structural and molecular underpinnings of physiological function and disease.
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Acknowledgements This work was funded by a National Institutes of Health (NIH) Director’s Transformative ResearchAward (TR01) to K.D. from NIMH, aswellas byNSF, the Simons Foundation, and the President and Provost of Stanford University. K.D. is also funded by NIDA, the DARPA REPAIR program, and the Wiegers, Snyder, Reeves, Gatsby, and Yu Foundations. K.C. is supported by the Burroughs Wellcome Fund Career Award at the ScientificInterface. S.-Y.K. is supported by a SamsungScholarship, A.S.A. by the Helen Hay Whitney Foundation, K.A.Z. and A.K.D. by an NSF Graduate Research Fellowship and J.M. by the NIH MSTP. We acknowledge H. Vogel, L. Luo, L. Schwarz, M. Monje, S. Hestrin and D. Castaneda for advice, and the Autism Tissue Program for providing human brain tissue, as well as J. J. Perrino, J. Mulholland and the Cell Sciences Imaging Facility at Stanford for electron microscopy imaging and advice. We would also like to thank the entire Deisseroth laboratory for discussions and support. CLARITY resources and protocols are freely supported online (http:// CLARITYresourcecenter.org).
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