I have NK-92 cells from ATCC. I used two different media for cell expansion.
One of them is CellGro-DC media with %5 FBS, %1 penicilin, 500 IU/ mL IL-2,
Other one RPMI 1640 media with %20 FBS, 2mM L-glutamine,%1 penicilin, 500 IU/ mL IL-2.
For the first stage of cell expansion I cultured cells at T-25 flasks in 10 mL medium with each media. However, 24 hours later , More than 90% of the cells were dead. Viability was 8%. I wonder why NK cells died.
Can anyone explain it and give me some advice?
Hi Dilara, why aren't you using the media recommended by ATCC?
https://www.atcc.org/Products/All/CRL-2407.aspx#culturemethod
I'll tag along with Steingrimur's question. I would give a closer look to your high concentration of IL-2 (2.5-5 times higher than the ATCC recommendation). Since the cytokine seems to play a potent role in the cells' activation or at least in increasing their acticity towards target cells it might be detrimental in the culture?
Hi Dilara, I agree with Angela, the IL-2 concentration you use is probably too high for NK-92 cells, since with just a few of it they are fine. Normally, one uses 500IU/ml of IL-2 for physiological human NK cells because they don´t grow so well. You are probably over-activating them, so they enter into apoptosis.
I would recommend you to use any concentration between 10- 100 IU/ml of IL-2.
I cultured them recently in TexMACS medium supplemented with 5% human AB serum and 100 U/ml IL-2 and they grow quite well. I didn´t add penicilin, neither the horse serum that ATCC recomends, and the cells were fine.
Thank you for your suggestions. I changed the media and cultured them in Alpha-Mem medium supplemented with %10 FBS, 2mM L-glutamine, %1 Penicillin 0,1 uM b-mercaptoethanol, 100 U/mL IL-2. Now they grow pretty well. It works after these changes. Thanks a lot.
We grew ours in Miltenyi MK-MACS media supplemented with 250 IU IL-2 and 5% heat-inactivated human AB serum.
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