I have been culturing 293T cells since last 2 years and never had this issue. They develop multiple vesicles seeding after 2-3 days, fail to grow and degenerate. I have tried changing media with fresh cells, but with same results. Are they infected or some other deficiency ? Kindly refer to the attached pics.
That sounds odds to me. When I was in Lu lab, I had similar experience that 293 T cells grew very slow (but they did not die). And then I figured out that the new coming-in medium was different from previously used; one component was not pre-added. If you get no answers on infection or contamination, I recommended to check different batches of cryo-stocks, check medium recipes and also dishes (not kidding, I found someone bring in petri dishes to culture one day).
Thanks Danyang, I think issue is with the media, I am not the only one who has this issue
thanks again
You can fairly easily check for contamination by growing your cells in media without antibiotics (if you use antibiotics). If nothing takes over after a few days, they are most likely not contaminated. You could also test for mycoplasma and see if that could be it. There are different methods for doing this and different kits out there. We use one from Lonza
Exosomes? Maybe you need to have new population of same cell type? there is aging in culture...why not? maybe the Hayflick curve is at its peak and there is a limit in cell life...???
Hi
Are you using FBS in your medium or not? It occurs when we do not use FBS or use free antibody FBS , another problem is pH of your medium ,please put your medium in co2 incubator befor using, it can set PH of your medium.
it is 10% FBS with 1% L-Glutamine, 1% Na Pyruvate and Pen Strep and we use 5%CO2
Hi!
It looks like nutrient deficiency. It happened to me when we changed the medium to a pyruvate-free, the cells would attach and fail to grow after 2 days (HT-29)
Glutamine isn't stable. Some cell lines don't mind it going off whereas others will grow poorly. How old is your media? If it is past 1 month it might be because they are sensitive to low level of glutamine. If you don't use your media up within 1 month, you could try to add glutamax instead of glutamine as this is a lot more stable.
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