I am trying to grow MCF 12A cell line using DMEM/F12 medium with all the hormonal composition except Cholera toxin. The cells are not growing well only a few were adhered and remaining died.
Is there any substitute for Cholera toxin or the whole media do I have to use other than DMEM/F12?
MCF-10 and MCF-12A cells will be grown in a 1:1 mixture of Dulbecco’s minimum essential medium eagles (DMEM)and Ham’s-F12 medium, 20 ng/ml epidermal growth factor, 100 ng/ml cholera toxin, 10 μg/ml insulin and 500 ng/ml hydrocortisone , supplemented with 10% heat-inactivated fetal calf serum (56°C, 30 min),100u/ml penicillin G, 100 μg/ml streptomycin and fungizone (250 μg/l) (penicillin, streptomycin and fungizone).....This is the media for my cells and never have a problem
MCF-10 and MCF-12A cells will be grown in a 1:1 mixture of Dulbecco’s minimum essential medium eagles (DMEM)and Ham’s-F12 medium, 20 ng/ml epidermal growth factor, 100 ng/ml cholera toxin, 10 μg/ml insulin and 500 ng/ml hydrocortisone , supplemented with 10% heat-inactivated fetal calf serum (56°C, 30 min),100u/ml penicillin G, 100 μg/ml streptomycin and fungizone (250 μg/l) (penicillin, streptomycin and fungizone).....This is the media for my cells and never have a problem
Thank you zohreh riahi, for the reply. I have all the media component you have suggested except 100 ng/ml cholera toxin in the media. Without this cholera toxin, my cells are not growing. I am using the pre-prepared mix of DMEM/F12 media. Is it the media or the cholera toxin which I am not adding in the media.
I will also be growing MCF-12a cells without cholera toxin available to me. Do I need to replace it with something else and/or change the quantities of the other agents in the medium?
After searching for the cholera toxin's mechanism of action on cell proliferation, there may be a reason to avoid using it on using cells like MCF-12a serving as non-transformed tissue-matched controls for transformed epithelial breast cancer cells (like MCF-7): cholera toxin changes mitochondrial ultrastructure from 'orthodox' to 'condensed', resulting in major metabolic alterations (transition from State 4 to State 3). This would appear to change the metabolic phenotype of a non-transformed cell closer to a transformed one (hence the increased ability to survive in culture and replicate fast)
http://link.springer.com/10.1007/BF00869240
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