Hello all
I'm working with SHSY5Y cells and I always made Cryo cells from them, them thawed them and used them without problem. But, recently, after thawing, they are remain floating and not attach to the flask. I've never seen this before. I always did the same procedure and it's first time happen.
The Cryo tubes are not kept for long time. Recently I made them.
I really appreciate if you can advise me to solve this problem.
Thank you.
Hello Elham Amini. E.A
Where have you stored the cells. Have you stored the cells at -80 degree C? If yes, for how long? Mammalian cells are not recommended to be stored at -80 degree C for long-term storage, instead you need to store these cells in liquid nitrogen.
Have you taken the viability count after thawing? This will give you an idea about the viability percent. If you have recorded a good viability of say about 85-95%, then you may have to consider other factors such as environmental stress for this type of a problem.
What could cause environmental stress?
You may check out for these.
1. Contamination,
2. Inappropriate cell surface for attachment,
3. Temperature fluctuation in the incubator,
4. Improper gas mixture in the environment,
5. Inadequate or altered nutrients in the culture media. Have you changed the FBS (either the brand or the lot)?
Best.
Hi Elham,
I would recommend to use different batches of cryotubes and flasks, so to exclude the possibility of an artifact in them. If the problem persists then something genetically would have happened, which would be very interesting. In that case, you would need to go back to your original cell culture for recultivation. Hope you solve the problem soon. Good luck!
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