Am culturing PII cell lines with a media containing ultra centrifuged (100,000g) Fetal Bovine Serum to get rid of the extracellular vesicles. every thing was fine at the binning I did not even see a trace of contamination. out of sudden I keep getting contamination, and whenever I use another media which dose not have this special serum every thing is fine and cells seems to grow just fine!!
At the beginning I thought maybe the media bottle was contaminated so I changed it with a new one, but the contamination did not go !! so I filtered the media and here comes the problem, the media in the flasks turned yellow and the flask is not even confluent!!! did anyone had a similar problem? am struggling with this nightmare for a month now!
Hi Amna Alrabeea,
My suggestion if for you to check the incubator and culture room. I had this problem before on my previous lab. We found out that the water in the incubator was contaminated. Now that you know that the Media is okay you may test the serum and incubators.
Good luck
Hello Amana
I had the same problem when I worked with cell line. The reason was contamination by mycoplasma in the old aliquots stored at -80ºC. It was a really nightmare. I advise you check it
Regards
Hello Amana,
Important question: do you routinely use antibiotics? (generally not recommended, as it can mask the contamination)
At first, I would recommend you to check the serum sterility - take new cultivation flask, add some media (without antibiotics!) and the serum. No cells. Leave the flask for few days in the incubator. If contamination appears, it is probably the serum.
Also it is important (as Erika mentioned) to keep the incubators clean. And you definitely should test your cells for mycoplasmas..
Hello Erika, Oscar and Jakub,,
Thank you all for your answers!!
Regarding the antibiotics Yes I do use them whenever I prepare a media, I will try all your advises today!! hoping that i would carry on my experiment without contamination.
Thank you very so much!!
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