Since 2 weeks I am trying religiously to revive my iPSC cell line ,but to my dismay the cells are not getting revived.This did not happened earlier.For the attachment of cells I am using vitronectin coating on tissue culture treated plates.The very next day of revival earlier used to show good results with growing cells ,but now the cells are not attaching and instead they are dead and floating.Basal mediam used is Esential8(DMEM ),but I am pondering over the change in colour of the media.The stored media bottles shows change in colour i.e,it has turned orangish instead of red.So should I check for the pH of the media.
Probably the media has changed pH, iPSCs are very sensitive. I rather recommended you to use new batch of media and try to cell counting prior to thawing or passaging.
pH is a good place to start with. Does this happen with a specific frozen batch of iPSC or any? You can also check your cell dissociation and freezing medium conditions.
Hello !! Muthu Kumar Krishnamoorthi Muthu Kumar Krishnmoorthi
Thankyou for your valuable suggestions Sir .I checked for the pH,it is between 7.3-7.8.So I guess there is no such issue with the pH.As you rightly pointed,there must be issue with the freezing(I use 10%DMSO).Even the time taken to transfer the cells from Mr.Frosty to Liquid Nitrogen or from Liquid Nitrogen to outside for revival might aafect.
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