I need to measure the GPF intensity in response to the pH change the issue I am facing is that the GFP is attached inside the cell and I do not know how to measure the intracellular pH with GFP intensity at the same time.
most of the pH indicators are dye I am afraid that that will overlap with GFP intensity
@Bertrand Cornu recommended I do that and quote ( One way to do this would be to use a pH-sensitive dye or probe, such as fluorescein, to measure the changes in pH in real-time. Fluorescein is a widely used pH indicator dye that exhibits a change in fluorescence intensity in response to changes in pH. To use fluorescein, you would need to introduce it into the cells. This can be done through co-transfection or by adding fluorescein to the culture media. Once the fluorescein is inside the cells, you can use a microscope equipped with a fluorescence detector to measure the changes in fluorescence intensity in response to action potentials.)
Howeover, how can measure the flourecent intensidty for co transfection both pH and GFP?
You'll need a fluorescent microscope that can differentiate different wavelengths. Typically, fluorescent microscopes come with 3 different filter sets: Blue (which is often used for Dapi), Green (used for GFP), and Red (used for RFP).
Then you can use a dye, as suggested, or a pH-sensitive biosensor based on RFP. For example, Tantama M, Hung YP, Yellen G. Imaging intracellular pH in live cells with a genetically encoded red fluorescent protein sensor. J Am Chem Soc 2011;133:10034-7.
No worries. Multi-color imaging is very common. GFP and RFP are quite different in wavelength and are easy to spectrally separate, i.e. they have little influence on each other. For more information I suggest you look at general information on fluorescent imaging. The free info from Addgene.com may be helpful. They have a "Fluorescent Proteins 101: A Desktop Resource" eBook.
Your major problem may be timing. Action potentials are usually quite fast, so your fluorescent proteins will need to respond rapidly and your microscope/camera will need to detect these rapid changes.
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