Q1. I am planning to use an ALP kit to detect the mES cells pluripotent/ differentiation status. Which would be the better (ALP) kit for this purpose?
Q2. I generally passage mES cells every 2-3 days. But some kit says 4-5 days (crucial for ALP activity). The detection days (4-5) are very strict? If grow the cells for 5 days, the cells were not looking good., may be started differentiated (EB?).
(I used 2i+LIF medium with supplements)
Q3. Does anybody having experience of using- Vector Blue Alkaline Phosphatase Substrate Kit III (from Vector Laboratories) for mES cells?
Thanks in advance!
Hi Ramanan,
I work with human iPSC and I guess 4-5 days could be sometimes too long for ESC which might lead to differentiation. Proliferation rate might differ from cell line to cell line (sometimes even for clonal lines), so I would recommend to passage your cells as you normally do (AP staining would not work when the cells are differentiated, right :)
I tried live AP staining on my iPSC, which worked nicely. One advantage is that you do not have to fix the cells and they survive the staining with no differentiation or some other side effect. Have a look at the link if that might be of interest to you.
Good luck.
https://www.thermofisher.com/order/catalog/product/A14353
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