Hello everyone
I'm working with ATCC-BYS0111 Human [Hispanic Male] Induced Pluripotent Stem (IPS) Cells
ACS-1025™ and the thing is that their morphology is changing a lot, I've been following the ATCC protocol EXACTLY as their website says, however I haven't been lucky, apparently.
It's only passage 3, and I have several vials frozen, however I'd like to figure out what's going on before to start working with a new vial.
Thanks in advance!
Hi! Do you have any images from the first passages you performed starting from the thawing? This way we can figure out whether they were pluripotent in the first place, or lost their pluripotency during subculturing. Without reading your script, I'd say those cells were MEF feeders. It is very unlikely for spontaneous differentiation as long as you supplement rock inh. every time you subculture, freeze or thaw this cell line. It'd be the best to ask ATCC. My general advice would be that you may speed up during subculturing. The more time they spend without being attached to the dish, the more they spontaneously differentiate, this includes the time it takes for you to perform the cell counting, cell detachment or freezing.
Hi!
I do have some pictures, I'll attach them.
I have only been using ROCK inhibitor after thawing the cells, not every time I change the medium.
Oh they apparently lost their identity as you culture them. You are not supposed to supplement ROCK inh. every time you perform media change, but it really helps to use during subculture (passaging, freezing etc.). Please note that Rock I. is only needed for single cell passaging. If cells lost pluripotency despite being passaged as clumps, then there is another problem that should be sorted out, like contamination, coating reagents etc. Best of luck!
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