Protocol for generating a 3D culture of epiblast stem cells
Year: 2024
Type: article
Abstract: Mouse gastrulation entails concomitant changes in cell fate, tissue shape, and embryo size. The use of a reproducible in vitro system is crucial for dissecting the mechanisms that coordinate these events. Here, we present a protocol for generating a 3D culture of epiblast stem cells (3D EpiSCs), which grow as epithelial spheroids mimicking key features of the gastrulating mouse embryonic epiblast. We describe steps for spheroid formation, growth, and passaging, followed by imaging or further downstream analyses. For complete details on the use and execution of this protocol, please refer to Sato et al. 1 • Steps to generate mouse embryonic stem cell-derived and epiblast-derived 3D EpiSCs • Instructions for dissociating and passaging spheroids • Guidance on plating 3D epiblast stem cells for immunostaining or other analyses Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Mouse gastrulation entails concomitant changes in cell fate, tissue shape, and embryo size. The use of a reproducible in vitro system is crucial for dissecting the mechanisms that coordinate these events. Here, we present a protocol for generating a 3D culture of epiblast stem cells (3D EpiSCs), which grow as epithelial spheroids mimicking key features of the gastrulating mouse embryonic epiblast. We describe steps for spheroid formation, growth, and passaging, followed by imaging or further downstream analyses.
Source: STAR Protocols
Institution: MRC Laboratory of Molecular Biology
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $1,500
OpenAlex ID: W4402923817