کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
2035413 | 1072171 | 2014 | 14 صفحه PDF | دانلود رایگان |
• PACT: passive tissue clearing and immunolabeling protocol for intact thick organs
• RIMS: compatible storage and imaging media preserves fluorescent markers over months
• Single-molecule FISH compatible 3D phenotyping enabled for thick tissue samples
• PARS: whole-body clearing and phenotyping compatible with endogenous fluorescence
SummaryUnderstanding 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.
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Journal: - Volume 158, Issue 4, 14 August 2014, Pages 945–958