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Cell. 2014 Aug 14;158(4):945-58. doi: 10.1016/j.cell.2014.07.017. Epub 2014 Jul 31.

Single-cell phenotyping within transparent intact tissue through whole-body clearing.

Author information

1
Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
2
Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA; Division of Dermatology, Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095, USA.
3
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
4
Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA. Electronic address: viviana@caltech.edu.

Abstract

Understanding the structure-function relationships at cellular, circuit, and organ-wide scale requires 3D anatomical and phenotypical maps, currently unavailable for many organs across species. At the root of this knowledge gap is the absence of a method that enables whole-organ imaging. Herein, we present techniques for tissue clearing in which whole organs and bodies are rendered macromolecule-permeable and optically transparent, thereby exposing their cellular structure with intact connectivity. We describe PACT (passive clarity technique), a protocol for passive tissue clearing and immunostaining of intact organs; RIMS (refractive index matching solution), a mounting media for imaging thick tissue; and PARS (perfusion-assisted agent release in situ), a method for whole-body clearing and immunolabeling. We show that in rodents PACT, RIMS, and PARS are compatible with endogenous-fluorescence, immunohistochemistry, RNA single-molecule FISH, long-term storage, and microscopy with cellular and subcellular resolution. These methods are applicable for high-resolution, high-content mapping and phenotyping of normal and pathological elements within intact organs and bodies.

PMID:
25088144
PMCID:
PMC4153367
DOI:
10.1016/j.cell.2014.07.017
[Indexed for MEDLINE]
Free PMC Article
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