Hi,
I am working on my PhD project and recently we plan to have our experiments using immortalized nasoepithelial cell line. First we tried ABM T9245 with the Prigrow serum the company suggested, with pre-coated vessels, however the cells didn't attach at all. For some other issue we asked for a compensating vial ABM T0440, along with the suggested serum and precoated culturing vessels. This is the second day after thawing, and the cells don't attach at all either.... Does anyone who worked with similar cell types have any idea what's wrong? or offer any suggestions?
I thawed the cells using 1 min 37 degree water bath, then resuspended in 5 mL Prigrow X, followed by 200g x 3 min centrifuge and finally resuspension and seeding as suggested density.
Hello Li-Fang Hsu , usually when adherent cells don't attach after the thawing it is because they are not well, but it can be for various reasons. How long have you kept them in culture to see if they could finally attach ? Some cells in very bad shape need more than 2 days to "heal".
Also, do you in your lab have any other way of thawing cells ? If yes, maybe you can try your usual method and see if there's any difference ?
The centrifuge speed seems a bit high for me (we usually do 1000 rpm for 10 min) but I don't think it would have such an impact on your cells.
Does your tube come from the cell bank, or from somewhere else (your lab's stocks or another lab's) ? I ask because the way they were frozen can greatly impact their shape after thawing ...
Hope this helps a bit, and good luck !
Hi, thanks for all the answers provided. I am here for some updates. Very small portion of the cultured cells survived after one week, and after "3 weeks" of culture it reached confluence. I passaged last week. These are the updated photos. To me its a lesson learned that some epi cells take so long to attach and survive...
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