[Cytometry] Compensation with liver cells

SIMON MONARD smonard at staffmail.ed.ac.uk
Fri Jul 10 05:24:34 EDT 2009

When you say beads which sort of beads are you using? I assume  
antibody capture beads, if so then the procedure is to set up voltages  
with unstained cells then do the color compensation with the beads.  
You will likely find them unusable as the commercial ones are designed  
for use with lymphocytes. If you are planning on doing much FACS then  
I suggest you make your own capture beads using beads with a  
background fluorescence comparable to your unstained cells. If you  
want to talk about this email me.


Quoting "Chaudhary, Kunal" <kchaudhary at ukaachen.de>:

> Dear fellows,
> I have a big problem regarding compensation. I did compensation with  
>  beads which was accurate but as soon as i tried to compensate with   
> liver cells its shows huge difference. I dont know which   
> compensation set up to take and which not. I isolate the liver cells  
>  by perfusion (Collagenase type 1 + Dnase I) and than digestion  
> again  with collagenase + DNase I at 37 degree. Finally i measure my  
> cells  and i see again wired results. I didnt add any Ab labelled  
> with  percpe Cy7 and APC but still i get strong signals. I rechecked  
> the  PMT and compensations and all but i still cannot understand  
> from  where i get these signals. Please please help me.
> Thank you very much in advance,
> With regards,
> Kunal Chaudhary
> PhD student
> Department of Medicine III
> University Hospital Aachen
> Aachen, GERMANY
> Tel: +49-241-80-89200
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Simon Monard
FACS Facility Manager
MRC Centre for Regenerative Medicine
Institute for Stem Cell Research
University of Edinburgh
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