Hi Kirk,
FCS Assistant can be used to generate ratio's, I understand that some
people are using it for indo 1, others use it to ratio Fluo-3 with Fura
red or SNARF. As far as validity goes, you've reminded me that I've been
meaning to get in touch with Jim Jett from Los Alomos, to request reprints
of two papers he referred to in a brief article in a copy of the NFCR
newsletter that I picked up at the last ISAC, which point out artefactual
fine-structure variations in ratioed digital measurements.
The references are:
Johnston,R.G., Schroder,S.D., and Mallawaaratchy, A.R., "Statistical
Artifacts in the Ratio of Discrete Quantities", The American Statistician,
vol 49, 285 (1995)
Jett, J.H., "Flow Cytometric Measurements of Cell Surface Phenomena", Acta
Path. Microbiol. Scan. Sect.A, Supp.274, 187-190 (1981).
But I don't have access to them locally (if Jim Jett is reading this could
he send me copies please?).
As far as I know, mean and median measures of calculated-ratio populations
are largely unaffected, as are the gross forms of plots derived from
calculated ratios.
If you are going to use FCS Assistant to generate your ratio values, be
sure to gate out (easiest on acquisition) events that have maximum values
for the parameters that you want to ratio, or these will give misleading
quotients (eg events that are in the top right corner will all ratio to
one, giving a spurious peak in the histogram). I didn't add a routine to
disregard these events, since not all cytometers use the full scale
resolution quoted in the file header $PnR value eg FACS Vantage and FACStar
plus parameters seem to have a maximum value of 1000, not 1023 as you'd
expect (which must be annoying for those who linearise log data - but
that's another story)
I've put some example figures on my server:
http://facsmac.med.cam.ac.uk/web/ratios.html
if you're interested
Ray
At 11:00 +0000 2/2/98, Kirk Watkins wrote:
>Is anyone using a calculated ratio for displaying Indo-1? The
>instrument does not have the ratio parameter but does have the optics,
>laser, etc. for running Indo-1. Can FCS Assistant be used to modify the
>data file to add the ratio parameter and is this valid for measurement?
>Any and all help would be appreciated.
>
>Kirk
Ray Hicks
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