[Cytometry] Quenching Autofluorescence

Karin Provost kprovost at buffalo.edu
Fri Apr 17 10:07:58 EDT 2009

I am looking for any suggestions on quenching red (and green) autofluorescence in
unfixed alveolar macrophages for multicolor flow cytometry. These cells
fluoresce bright green under a standard microscope and very brightly in all
channels of our cytometer (fitc, pe, pecy7, percp cy 5.5, apc, apc750).
We have tried the following protocols, with the outcomes listed. 

Have tried:
B-D galactopyranoside (20% reduction but still very bright)
Crystal violet (decreased FITC/PE but increased in all red channels)
sodium borohydride
Tween 20
nonfat milk
combinations of all of the above

Any thoughts on pH alterations, Janus dyes, light bleaching or other options?
Many thanks-

Karin Provost, DO
Research Assistant Professor
SUNY at Buffalo
School of Medicine and Biomedical Sciences 
VA Western NY Healthcare System
3495 Bailey Avenue
Room 823B1
Buffalo, NY 14215
(716) 862-7378

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