I see its also a common practice when labelling with TMT and SILAC. Also, would culturing in FBS-free media be preferable since serum proteins found in FBS may theoretically be detected when undergoing MS/MS?
I culture in phenol red for SILAC experiments. There is no chemical reason that TMT or SILAC would be incompatible with phenol red. Can you please provide more information or citations for this "common practice?"
Hi Boris,
I agree with Jesse, I have never seen anything about not using phenol red prior to iTRAQ labelling (or TMT, or SILAC).
As for FBS-free media, it all depends on your sample preparation, but I have never found it a problem. Even if you do detect them, why would that be a problem? As long as they don't dominate the signal, it should be fine.
Hi Boris
I cannot comment on your particular system but generally speaking if use of phenol red is contra indicated ts because phenol red can act as an agonist of certain cell receptors; in particular oestrogen receptors as I recall. Thus, for those subset of cell lines growth and biochemistry can be affected distorting studies. This applies if I recall correctly to certain metastatic breast lines
Hi all,
Thank you for your input. After reviewing the literature, it seems that nearly all the studies using phenol red-free media were conducted in breast cancer cells to account for possible estrogenic activity, or were interested in secreted proteins rather than the intracellular proteome. Since I am interested in neither, I will continue with phenol red.
Thanks all
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