Consider the animated picture to the left. It shows three populations of beads, one unlabelled and one each labelled with FITC and PE. Watch as the red and green coloured populations
move from an uncompensated to a correctly compensated, then an over compensated position relative to the X and Y axes.
So when is a population correctly compensated?
The short answer is
when the median of a population positively labelled for Y is the same as the median of a population negatively labelled for Y when displayed relative to the X axis.
Now let's use the simulator to adjust for spectral overlap on some real data files. The simulator will work with any FCS compliant data file, so you can practice and become comfortable with this concept using any file in the database, however if you'd like to use some data files which contain completely uncompensated data please select the following files:
Unstained:
FITC only:
PE only:
PerCP only:
APC only:
The basic procedure to follow is: