Question
Asked 22nd Apr, 2021

Why does the amount of rhodamine b fluorescent dye decrease so much when I wash the emitted cell with PBS?

when I wash the cells that emit fluorescent by PBS for flowcytometry, emittion of rhodamine b is very low

All Answers (3)

22nd Apr, 2021
Adam B Shapiro
Entasis therapeutics
The cells may have one or more efflux pumps (such as P-gp and MRP) that can pump the dye out. The function of these pumps can be blocked by treatments that de-energize the cells. Additionally, dye that is simply bound to the surface may be washed off.
26th Apr, 2021
Chathura S. Abeywickrama
St. Jude Children's Research Hospital
May be your dye has not been internalized in to the cell and retaining in the surface as an aggregate. Probably you may need to double check your staining protocol and provide sufficient time for the internalization. I would highly recommend you to try the same dye/method in a different cell line as a control. Hope this help!
4th May, 2021
Mª Angeles Zorrilla Lopez-Perea
University of Malaga
I am not sure if rhodamine b are sensitive to substances like washing, buffer or even PBS. Make sure the fluorophore is hold tight on the sample.
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