Fast and high-fidelity in situ 3D imaging protocol for stem cells and niche components for mouse organs and tissues

• Published in: STAR protocols 2022-09, Vol.3 (3), p.101483-101483, Article 101483
• Main Authors: Saçma, Mehmet, Matteini, Francesca, Mulaw, Medhanie A., Hageb, Ali, Bogeska, Ruzhica, Sakk, Vadim, Vollmer, Angelika, Marka, Gina, Soller, Karin, Milsom, Michael D., Florian, Maria Carolina, Geiger, Hartmut
• Format: Article
• Language: English
• Subjects: Cancer; Cell Biology; Microscopy; Protocol; Stem Cells
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2022.101483

Quantitative 3D imaging of organ-wide cellular and subcellular components is central for revealing and understanding complex interactions between stem cells and their microenvironment. Here, we present a gentle but fast whole-mount immunofluorescence staining protocol for 3D confocal microscopy (iFAST3D) that preserves the 3D structure of the entire tissue and that of subcellular structures with high fidelity. The iFAST3D protocol enables reproducible and high-resolution 3D imaging of stem cells and various niche components for many mouse organs and tissues. For complete details on the use and execution of this protocol, please refer to Saçma et al. (2019). [Display omitted] •Whole-mount immunostaining protocol for 3D microscopy of stem cells and niche components•Fast pipeline for multiple mouse organs and tissues•High preservation of mouse tissue morphology and molecular integrity•Allows long-term (up to 5 years) storage of samples for subsequent staining and imaging Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Quantitative 3D imaging of organ-wide cellular and subcellular components is central for revealing and understanding complex interactions between stem cells and their microenvironment. Here, we present a gentle but fast whole-mount immunofluorescence staining protocol for 3D confocal microscopy (iFAST3D) that preserves the 3D structure of the entire tissue and that of subcellular structures with high fidelity. The iFAST3D protocol enables reproducible and high-resolution 3D imaging of stem cells and various niche components for many mouse organs and tissues.