We ordered MCF10A cell line from ATCC and seed the cells in suggested conditions except adding cholera toxin and pen/strep. We made three stocks in this conditions but then suddenly our cells started to die and we cant stop it.
Our conditions are NOW : MEBM medium + MEGM kit + 100 ng/ml cholera toxin.
We tried to remove cholera toxin but the results are same.
Can anybody tell me where we are wrong?
Thank you.
I have also used MCF10A 3-D culture in Matrigel after reading the top-journal papers published from Professor Joan S. Brugge's lab. I attach the file about the detailed information about the culture medium.
In my opinion, the concetration of EGF ligand in the medium or the contamination with Mycoplasma might be responsible.
http://brugge.med.harvard.edu/protocols
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