Hi guys! I have a question regarding my CCD-18co cells. They grow very slowly and it is written on ATCC website, that "Growth of the cells is enhanced by addition of tumor necrosis factor alpha (TNF alpha) to the medium". What should be the concentration of TNFalpha? do I add it once in the culture media or should it be added fresh all the time ? should it be human recombinant TNFalpha from the yeast or E. coli? Thanks in advance!
Hi Dear Maria
I would suggest you to use a dose of 100pg/ml to your culture. You can use bacterial purified TNF which is generally active, if you wanna make sure about the activity of your TNF you can check the activityby treating L929 cell line which is susceptible to TNF (10ng/ml 48 h) and see the cell death, and cell death can be measured by MTT. most probably your TNF activates JNK and promoting cJun towards cell growth. I suggest you to add TNF when you change the media.
100 pg/mL of TNF sounds like a lot; I could be wrong since I have never worked with this cell line. Cytokines are usually added in the ng range when working with immune cells (which I have experience with).This is the paper that ATCC refers to. You could maybe follow what they did.
http://science.sciencemag.org/content/230/4728/943
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