We are trying to grow LUHMES cells. The cells attach well and look good for upto two days, However when we do a media change they end up dead on the next day. We checked all our media components (DMEM/F12, N2 suppliment, bFGF). Points already taken into consideration are: We have other cells growing perfectly well in our incubator, indicating no problem with the CO2 levels. We prewarmed the media at 37C. Added fresh bFGF. All our media components and coating reagents are new. Coating was done with Poly L ornithine and fibronectin as mentioned in ATCC. Please help!
Hi,
I have recently come across this issue too. I think passage number is a factor, LUHMEs don't behave well beyond passage 6. Another factor is oxidative stress. If you have HEPES in your media, ensure that it has not degenerated due to light exposure. HEPES is broken down into hydrogen peroxide due to photo-oxidation, and LUHMEs are extremely sensitive to any pH changes or oxidative stress. I know this question was asked a long time ago, so if you were able to solve the issue please do let me know!
Hi,
I have the same issue and would greatly appreciate any tips possible. My observations are that they have a fixed half-life and do not go beyond passage 11-12. It did help to use fresh media and freshly added L-Gln. But I still have to thaw new aliquots every month.
I see many aggregate cells forming ball-like in the growth media. Why is this happening?
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