Hi everyone,
I have been trying to differentiate hiPSC into cardiomyocytes. Once the beating of one well reaches about 50-70%, I replate them into different well plates for my further experiments. However, after staining I noticed that the purity of the cardiomyocytes was not good. Before "the actual" replating of cells for experiments, do you suggest any other replating or is there any way to expand hiPSC-CMs?
Thanks in advance!
Zeynep Elif Yesilyurt please have a look at this STAR protocol if you would like to expand CM numbers. Article Massive expansion and cryopreservation of functional human i...
However if you want to improve the purity of your CM pool then you can consider using different methods such as including a metabolic selection (Article Distinct Metabolic Flow Enables Large-Scale Purification of ...
) or purifying /sorting cells in case you have an endogenous fusion protein in your cell line.
Hi,
Not every differentiation of hiPS cells leads to obtaining a pure population of differentiated cells. In the case of low differentiation efficiency, differentiated cells enrichment is necessary.
In the case of differentiation of hiPSC cells into cardiomyocytes, it is recommended.
I suggest using immunomagnetic sorting: EasySep™ Human PSC-Derived Cardiomyocyte Enrichment Kit.
The EasySep™ Human PSC-Derived Cardiomyocyte Enrichment Kit is designed to isolate human PSC-derived cardiomyocytes from cell cultures by negative selection. Unwanted cells are targeted for removal with Tetrameric Antibody Complexes recognizing non-cardiomyocytes and dextran-coated magnetic particles. The labeled cells are separated using an EasySep™ magnet without the use of columns. Desired cells are poured off into a new tube.
https://www.stemcell.com/products/easysep-human-psc-derived-cardiomyocyte-enrichment-kit.html
Good luck !
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