Protocols for differentiation of pluripotent stem cells into specific populations e.g. neurons or hepatocyte-like cells have been reported. We have optimised these protocols by implementing magnetic cell sorting steps.
Magnetic enrichment of pluripotent human iPS cells prior to differentiation leads to more predictable differentiation results and allows automation of the workflow.
Generation of Hepatocyte-like cells (HLC) in vitro is usually achieved by exposing PSC to high doses of activin A in combination with FGF and Wnt pathway activation. This leads to induction of nascent definitive endoderm (DE) that can be subsequently specified to hepatic endoderm (HE), hepatoblasts, and finally metabolically active HLC.
However, varying efficiencies of PSC differentiation are well documented and caused, e.g., by cell line predisposition, lot-to-lo variances of differentiation media components, and experimental inconsistencies. To overcome these limitations, we sought to develop an immunomagnetic selection strategy for the isolation of DE cells, thereby standardizing procedures for the differentiation of the endodermal lineage.
We see that isolation of DE cells enables reproducible generation of highly enriched HLC cultures and compensates for inherent variabilities during PSC differentiation, resulting in a more standardized differentiation procedure. The protocol may also serve to improve generation of other cell types derived from DE, such as pancreatic beta cells or lung epithelial cells.
Moreover the concept of enriching certain cell types during the differentiation process can improve the differentiation into other linages as well.