Although I can't answer the motivations of people, one generally wants to measure P (phosphorylated) ERK, but PCR based techniques of course look at RNA levels, which don't have perfect correlation with protein levels. The protein levels are not the phosphorylated protein levels, which people interested in P-ERK are looking at.

Immunohistochemistry is generally used when you have tissue slices, in which you can see the localisation of the antibodies. This allows for semi-quantification while not having to fully mix your cells. Western blot quantifies protein, but the original localisation will be unknown and be partly based on how you separated your tissue beforehand.

I like to note that I personally don't see much use of doing this for cell cultures, but I hope that others may shed more light on this or correct me on the above.

Goodluck!

I agree with Jelle-

To see phospho-ERK you will need to do a western blott with an antibody specific for p-ERK and to harvest your samples with a phospho-buffer in order to preserve the phosphorylation status of your protein/ERK

You may be able to perform IHC for phospho-ERK but you again will need to prepare your samples in such a way to preserve phosphorylation status

PCR will not allow you to determine the amount of phospho-ERK protein as PCR assess the amount of RNA present and not actual amounts of total protein or phospho-protein. ERK function is regulated b phosphorylation so looking at levels of phospho-ERK typically provides a more meaningful idea on the activation of the p-ERK pathway.

cFos is one component of a transcription factor complex- cFos typically heterodimerizes with cJun to form AP-1. The activity of cFos depends on nuclear translocation- consequently you need to use a tecnique that can determine if cFos (and its dimerization partner) are in the nucleus. To do this you need to look at nuclear localization which can be determine via IHC or by western if you first perfomr a nuclear extraction kit.

Thanks, Jessica Mandula &Jelle Bonthuis