I tried for the first time to see the fluorescence of a drug, when it is internalized into MCF-7 cells.
But I have observed a really bold, endogenous fluorescence in my control cells.
What could I possibly do to overcome this obstacle?
Thank you in advance for your answers!
Intrinsic Fluorescence from cells is very common. There are many proteins containing aromatic amino acids or highly charged amino acid residue and endogenous pigment which give fluorescence in wide range (mostly emits in blue/ green spectrum). So, try some following points
1. Choose the fluorophore which emit in red region such as CoraLite594 and CoraLite 647, will help distinguish specific staining from Intrinsic fluorescence from native compounds.
2. Treatment with sodium borohydride (NaBH4; 0.1–2.0% in Phosphate Buffer).
3. Use the filters which helps a lot without adversely affecting the cells; Here some filter available:
chromatic beamsplitter FT 510, bandpassfilter BP 515–565 for argon laser irradiation at 488 nm and FT 560 longpass filter LP 570 for helium neon laser irradiation at 543 nm.
I hope it will help.
Regards
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