Technical Question....

Leary, James F. jleary at utmb.edu
Wed Aug 5 17:40:05 EST 1998


On Tuesday, August 4, 1998 David Burke asked the question:

 
I recall some time ago that the number of positive events detected by
flow cytometry does not directly correlate with actual number of cells.
For instance, if 50,000 positive events are detected, can one accurately
take that as 50,000 CELLS?
 
If not, what exactly is the deviation and how can the degree of
deviation be calculated???
 
Any references or helpful inputs????
 
Thanks


David:

The question has a number of facets, some involving sampling statistics,
but in the case you are probably interested a more important factor is
the instrument deadtime.  The instrument's ability to see cells and
count them accurately is a function of all of the deadtimes of the
various parts of the instrument as well as the ability of the instrument
to resolve two cells as two and not one.  I spent a fair amount of time
discussing these issues and providing sample calculations (so you can
easily do your own calculations or even easily determine the deadtime of
your instrument) in a book chapter I wrote back a few years ago. I also
provide graphs so you can visualize the effects of various factors on
this deadtime problem. A large number of people have found it pretty
readable and helpful to them. The reference is: Methods in Cell Biology:
Flow Cytometry, vol. 42, chapter 20: "Strategies for Rare Cell Detection
and Isolation", 1994. Probably less of interest to you, but a paper we
wrote dealing with the sampling statistics side of the problem was
published last year in Cytometry 27: 233-238, 1997: "Theoretical Basis
for Sampling Statistics Useful for Detecting and Isolating Rare Cells
Using Flow Cytometry and Cell Sorting".  Good luck!   -- Jim Leary



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