GFP western - (Feb/20/2008 )

Hey ,
So I have two sets of cells. One set expressed GFP 2x the second set (via microplate reader). Will I be able to detect the difference in GFP levels using a western? IF not any have suggestions on how I can show a two fold difference in gfp expression?
Thanks
joe

You could detect the difference in expression using GFP antibody on Western..

I don't think western blots are very quantitative. They could give you a rough estimate sometimes but often it depends on the level of saturation: that is ,your blot will show differences only if the expression levels of GFP in both lines are low. If both lines express high levels of GFP, you may have to load very little protein, use little antibody and expose for a short time to show up differences- even then it is not quantitative. I would advise doing a northern blot or real time PCR for accurate fold difference measurement of GFP transcript levels. Alternatively, you could do FACS(flow cytometry), which will give you the mean fluorescence levels for your two sets of cells.