log-fsc x log-ssc vs lin-fsc x lin-fsc and FL1

Gerhard Nebe-von-Caron Gerhard.Nebe-von-Caron at Unilever.com
Wed Aug 26 11:04:22 EST 1998



          Agreeing strongly with Ray's comment about the thresholding 
          exercise.  When you want to convert to log scatter data 
          display it is a good idea to display log and lin side 
          scatter versus fluorescence in two pictures side by side as 
          I do on our Coulter XL. Thus when triggering on the linear 
          signals you can familiarise yourself with the position of 
          the clusters in log space and find the appropriate 
          discriminator level on the log detector.
          
          From the older systems I remember that there could be timing 
          problem when triggering on a log signal that could lead to 
          missing the fluorescence signal.  I do not know how the 
          timing works in the Calibur but would assume that less 
          likely to occur with modern electronics, but if in doubt 
          whether the cells should be fluorescent or not, just take a 
          look down the good old microscope.
          
          All the best
          
          Gerhard


______________________________ Reply Separator _________________________________
Subject: log-fsc x log-ssc vs lin-fsc x lin-fsc and FL1
Author:  janssen at Moffitt.usf.edu at INTERNET
Date:    21/08/1998 00:23


We have just completed, using a Facscalibur, analysis of the presence of
a genetically engineered surface marker on renal carcinoma cells.

Because a log-ssc x log-fsc representation gave a nice cluster of
events, we used those scales to acquire. But to our dismay, when
examining the FL1 histogram, it appeared that no cells were expressing
the protein being stained for. Subsequent acquisition using linear
scales for both fsc and ssc, showed substantial staining on the same
cells.

We are at a loss to account for why this should be so. Anybody out there
have an idea? Anybody at B-D want to explain?

Thnx Vry Mch!!!

********************************************************************
William E. Janssen, Ph.D.
Associate Professor
University of South Florida / H. Lee Moffitt Cancer Center
12902 Magnolia Drive, Tampa, FL 33612, U.S.A.
phone: 1-813-979-7202; fax 1-813-979-3071



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