This content is Free Access.
Chapters
- 00:05Title
- 03:34Generating Conditioned Medium (CM) For the Colony Assay
- 05:13Obtaining a Single Cell Suspension of Pancreatic-like Progenitors by Fluorescence Activated Cell Sorting
- 07:55Plating Sorted Cells to Semi-solid Culture
- 10:49Observing Colony Formation under the Light Microscope
- 11:47Ngn3-EGFP+ Derived Colonies Express Insulin
- 13:21Conclusion
A three-dimensional clonogenic assay that allows pancreatic-like progenitors to differentiate into insulin-expressing colonies is described. This method takes advantage of semi-solid media containing methylcellulose, Matrigel and growth factors, in which single progenitors proliferate and differentiate in vitro, permitting quantification of the number of functional progenitors in a population.
Tags
Quantitative AssayInsulin-expressing Colony-forming ProgenitorsPancreatic Stem CellsProgenitor Cell BiologyIn Vitro AssaySingle Cell AnalysisLineage PotentialPancreatospheresSuspension CultureSingle Cell DepositionCulture MediaNumerationLabor-intensivePancreatosphere-initiating ProgenitorsProliferation And DifferentiationSemi-solid Media Based Colony AssayMethylcelluloseThree-dimensional Space
