My DMEM has 3.7g/L of Sodium bicarbonate, instead of 1.5g/L (I only have this DMEM), suggesting that my sensitive endothelial cells need 10% CO2 incubator. However I am working in an academic institution where the incubators are set at 5% CO2 for everyone. Therefore I want to add HEPES in my DMEM instead, and put in the 5% CO2 incubator. Do you think that this is a good idea?
Hello Nurul,
Am I guessing correctly that it is still the b.End3 that are troubling you? According to Helms et al. (see attached link) it is possible to add HEPES to acquire a more robust and stable pH for your culture:
"TES, MOPS, or HEPES is added to the differentiation medium to obtain a high buffer capacity, which has shown to promote high paracellular tightness in the coculture models [17]. Also, this medium maintains a relatively stable pH when removed from the incubator, whereby experiments on live cells can be performed outside the incubator without a prior change of medium."
The recipe for their DMEM suppl. is added in the article I believe. They are using it for primary cells, so it should definitely not be more troublesome for the b.End3. I do believe, however that 10% CO2 might still be necessary if you don't want to force your cells into a new phenotype..
Good luck!
http://link.springer.com/article/10.1208/s12248-014-9628-1/fulltext.html
Ask a colleague for bicarbonate-free media and mix with your DMEM to achieve your goal
Dear Anette - Yes, correct. Thank you for your help!
Dear Michael - That is actually a good idea. But sadly, others are using EMEM.
HUVECs will tolerate a 50:50 EMEM/DMEM mix if your serum is of good quality.
Anal Biochem. 2014 May 1; 452: 19–24. doi:10.1016/j.ab.2014.02.015
See this for primary HUVEC
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