My lab ordered a vial of the DAUDI cell line, but we are facing issues as the cells are growing very slowly, even after 4 months. Additionally, most of them are dying after only 4 passages, whereas the company who sold us these cells had mentioned that they should live up to 20 passages. I would be extremely grateful if anyone could help me with the best protocol to expand the cells and provide some tips.
The cell density is important, you should seed DAUDI at 0.5-1 x 106 cells/ml; and maintain the density when splitting; cell culture may be started in 24-well-plates and later transfered to flasks; split ratio 1:2 to 1:4 every 2-4 days;
the culture medium should be 90% RPMI 1640 + 10% FBS; you may also increase FBS to 20%.
Dear Sabine,
Thank you for your answer and advice. We initially used 90% RPMI 1640 and 10% FBS, and split the cells 1:2 1.4. However, the cells were not growing, so we left them in the media for longer. Is it possible that they needed more fresh media, and that's why they didn't expand as expected?? And, do you always count the cells before dividing them?
It's very strange that under these conditions your DAUDI cells are not growing. Even if the cells are not properly growing, I would suggest to change the medium at least once a week; and I would increase the FBS to 20% to see whether this increases the growth rate, if it does, you can later on again reduce it to 10%. As long as the cells are not proliferating, I would not split them at all, just change the medium.
Are your cells still viable and just not growing or already dead?
Did you test the cells for mycoplasma contamination?
Can you post a picture of your cells?
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