Dear colleagues,
Thank you for reading my questions.
I’m currently culturing my patient derived iPSCs, and I have some problems these days.
The problems are:
1. Especially after media changing, cells suddenly are detached and died.
2. Cells morphologies are weird: the center of colonies are loosely packed and they do not have round shape.(passaging number is early: 7~10)
My iPSCs culture routine as follows.
1. Coat cell culture plate at least 1h in 37’C, 5% CO2 incubator, with Matrigel+DMEM F-12(1:100 working solution).
2. When I do passaging,,add 10uM of Rho kinase inhibitor in mTeSR plus media, and remove Ri following day. After that, i replace media every other day until the next passaging day.
3. 'Gentle cell dissociation reagent' was used.
I've been culturing ipsc with the same process for a few months, but I've recently encountered the above problems. I don't know which part is causing the problem. I need your opinion.
Thanks.
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