We are doing some sequential multiplexed IF experiments but are seeing that the emission spectra for the dyes is significantly changing (20-30nm red shift). Thus far we cannot explain it, other than perhaps it relates to antifade mounting media (although trying different types has not helped). Has anyone encountered this? Any suggestions about trying to avoid the phenomenon?
Jesica Canizo
when you make a multiplexed IF, fluorophore red shift its emission compare to its emission spectrum alone?
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Chris Carey
well it's odd. All of our dyes are moved 20-30nm red-shifted, whether in the multiplexed or single stained controls. We suspect it is down to the mounting medium and I wondered if that is a recognised phenomenon.
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