I have generated iPSC colonies from peripheral blood, today is Day14 post-transduction and the colonies seem to have acquired nice borders and distinct iPSC morphology (images attached). There is some difference between the different colonies in the same well, some are quite large while others are still small without defined borders. There are almost 45-50 colonies.
Is it better to passage them now using ReLeSR or should I pick them manually single colony-wise?
Hi there!
I'd suggest you perform AP live staining (SSEA4 or TRA-1-60 would also do) and pick at least 10 positive colonies from there manually. The remaining colonies can be enzymatically passaged. If you are aiming for clonal expansion, surely proceed with manual passaging.
As far as I can see from the images, not all the cells are fully reprogrammed within a colony, so dissecting those under microscope manually would help the pluripotent pieces to form independent colonies. For manual passaging, I'd also recommend using rock inhibitor if the sample is so precious for the initial passaging. Good luck!
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