Hi All,

I am trying to study FRET- live cell imaging with CFP-YFP pair targeted for PM and cytosol in HEK 293T cells. I want to know what settings or parameters should we have to monitor FRET. I am using sensitized emission method and have 3 channels which are must for that. I can see signals from all channels, but the problem is I am not seeing changes in the FRET signals upon stimulation (as expected and previously tested). Is there any specific setting needed so that I can use the in-built FRET analysis mode on Zen black? I only want to see increase FRET upon stimulation that I am not able to. A help and any suggestion or detailed protocol will be highly appreciated. Thank you