Hello Amudhan,

HEPES is a buffer to maintain the pH of the culture medium. it can be very important if you do not using a CO2 incubator, or if you are manipulating the cultures a lot, which brings them out of the CO2 atmosphere of the incubator.  Some cells/assay are more sensitive to fluctuations in pH than others, so in in doubt, it may be better to include HEPES as there are multiple cell types in PBMC cultures.  When I have used HEPES previously (not for PBMC though), we used a final concentration of HEPES of 10 mM (Ulaeto at. al. Journal of General Virology (1995), 76, 103-111).

Hope this helps,

David

Thanks David.

We will try with HEPES buffered media for our culture

Hi Amudhan, 

addition of HEPES is specifically required for pH sensitive cells like macrophage/monocytes. As PMBC contains these cells, so it might be better to add HEPES at a final conc. of 10 mM.

Thanks Deb.