I had stored HeLa cells in -80C freezer for over a month. When I thawed and seeded the cells today, I noticed that the cells looked significantly different than usual cells when floating in media. After 2hrs, I checked the cells again to see any changes in attachment behavior and found that the cells look different than usual in the attached condition also. I have attached pictures of the floating and attached cells. Please check and let me know if you could think of any possible explanation for this. I also observed similar behavior in DU145 cells that I thawed after a month. I am also attaching pictures of DU145 (less confluent) after three days of seeding.
Dear priyanka
Chnage in cell morphology is not uncommon, however it is very important to know the cause behind the change. Sometimes, we observe a sudden change in mrophology of our runing cultures(which usually changes back to normal after a passage or so), however this change owes to various environmental changes like change in incubator temperature, co2 percentage, humidity or cell health even the change could be due to some biologic contaminiation. So in that case we must examine these variables one by one to ensure the actual cause. However, on reviving your cryostocks, a change in morphology can happen but after a couple of passages the cells retain to their normal morphology. If you don't find cells retaining normal morphology, then chances can be that your cell could be cross contaminated which has the least chance but we must consider it too.
So in short I would suggest to wait for a couple of passages and see if the cells retain back to normal.
Good luck....
Rayees Padder Thank you, sir. The cells seem to be healthy after two passages now.
Hi Priyanka,
Have you thawed the cell directly from -80 where you stored them?
if so then the cells will not retain the same features on long term storage in -80 freezer. either you need to shift the cells to liquid nitrogen with in 48 hrs of storage to -80 freezer to maintain the cell health and original morphology. otherwise the cells will lose the normal morphology with long term storage in -80 and reviving them directly from -80 will not give the same results as you have revived them from liquid nitrogen.
so now you can understand from where you are thawing the cells and the reason for your problem.
They looked normal to me. After thawing, mine looked the same. So how do they normally look like after being revived?
Your cells will revert to normal morphology after a couple of passages. Such changes in morphology is common is cell lines revived from cryo-storage.
As Akhil said, cells tend to change morphology after thawing and they need just some time/ passages to recover. In my case, I work with human fibroblast and they can take up to a week to "wake up" after seeding.
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