Structural and Molecular Interrogation of Intact Biological Systems

Stanford Researchers Utilise the Leica TCS SP5 Confocal for Their "See Through Brain Studies"

April 12, 2013

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.

Read full article:

Kwanghun Chung et al.
Structural and molecular interrogation of intact biological systems

Nature 2013, doi: 10.1038/nature12107

The Nature Video reveals how Karl Deisseroth and his team created 3D visualizations of mouse brains

YouTube video by nature video

To understand structure and function of brains or other complex biological systems, the method of  choice is microscopy. In particular, confocal microscopy is employed to reveal three-dimensional connectivity and functional interactions. To come to a real insight into brain’s way of working, one must look deep into the tissue – which usually is non-transparent. A couple of clearing methods have been developed in the past, but they usually come along with distortions of the structures, incompatibilities with fluorescence stainings or are just prohibitively toxic to the lab technician.

Karl Deisseroth’s group, known for pioneering optogenetics, developed a new method to clear tissues that combines a set of promising features. The method, called CLARITY keeps the three-dimensional arrangement of biological structures from long neuronal connections down to the level of sub-synaptic patterns. The product of the treatment is transparent and permeable for macromolecules – enabling intact-tissue in situ hybridization, immunohistochemistry with multiple rounds of staining and de-staining in non-sectioned tissue, and antibody labeling throughout the intact adult mouse brain. It also 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.

CLARITY Protocol and Materials

http://clarityresourcecenter.org/

More videos with Karl Deisseroth

CLARITY process: interview with Karl Deisseroth at Stanford University

YouTube video by stanfordmedicine

Controlling the brain with light

YouTube video by StandfordUniversity

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