Should I still add 10 % Calf serum if I supplement the cell culture medium with 25mM HEPES (for incubation without CO2)?
I want to add HEPES as I dont have CO2 incubator
My culture medium originally contains: DMEM, 10% calf serum, 1% sodium pyruvate, 1% Pen/strep/glut
No. If your media already contains 10% calf serum you don't need to add extra. But it also depends on cell type. I have observed for certain cells (tenocytes), some people tend to add 20% calf serum to the media.
Hi,
HEPES and serum have different roles in culture medium. HEPES helps maintain the pH whereas serum adds nutrients, growth hormones etc.
Keep the orginial medium and just add the HEPES.
Hi dear Emily
Based on your cell line, the amount of serum will change.
I suggest you check your cell line at https://www.atcc.org/.
you can find all information about your cell line.
good luck
You need to check how much serum is required for the growth of your cell line/cells. Already your medium contains 10 % calf serum, if your cell line requires more, you have to add or else no need. Also, the function of HEPES and serum are different. You have to add both.
Of cours serum is required as it provides growth factors. Hepes only maintain the pH and replace the CO2/bicarb buffer. Caution some cells do not like Hepes.
HEPES is just an alternative for CO2 to maintain the pH. Also, check the media composition if it already contains serum then no need to add it additionaly but if its absent then you need to provide another source of nutrients for the cell like FBS 10%.
Dear Emily,
As other researches have pointed out, if your culture medium already contains serum, it would not be necessary (or even recommended) to add an extra dose to your wells/flasks. Indeed, the fact of adding serum or not is independent of the role of HEPES, which helps to maintain the pH. If viability becomes an issue, you really might want to try a CO2 incubator since most primary cultures and cell lines progress nicely well in 5% (v/V) CO2 (although some cells such as adult neural stem cells require levels up to 8% in adherent cultures), without leaving HEPES. Regarding the serum, I am fully aware its use is highly widespread, but if you are planning to study cell signalling, I would recommend you to check first if the pathways activated by the growth hormones present in it are not known to interact with the ones you want to examine, which could bias your results when trying to determine the specific and autonomous nature of those routes.
Hope this could be somehow helpful and good luck in your experiments!
Hi,
HEPES is a buffer which helps to keep the PH value. but serum is common supplement that helps cells to grow.
Due to the different roles and effects of HEPES and serum (as other researchers have noted) serum should not be removed from the culture medium.
Your comment appears like you are adding HEPES to the cell culture without any growth medium, if this is the case; firstly select a suitable growth
medium for the cell lines then adjust the pH with HEPES.