Re: gating on histogram overlap
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From: Maciej Simm <simm@treestar.com>
Date: Mon Oct 09 2006 - 14:34:25 EDT

Stevan,

the cytometry archives list a fair amount of discussions about
isotype vs. FMO controls, and I think in the case of dim antigens,
FMO will be a better benchmark for gating. I suggest reading up on
FMO's :)

You could try to measure the fluorescence intensity of your entire
histogram (ungated) which will probably show too small a change in
relative fluorescence to yield any interesting stats.

Maciej Simm
Tree Star Inc.
************************
Daily Dongle blog - http://flowjo.typepad.com/

On Oct 5, 2006, at 2:49 PM, Stevan Lauriault wrote:

Dear All,

When analyzing histograms of dimly fluorescent samples, and
comparing
to their specific
isotype controls on overlay, there is a significantly large overlap
between the two
populations (ex. ~10%).  Is there a universally accepted method to
statistically quantify
the relative proportions of positive vs. negative populations?

We are unsure whether to gate the entire histogram, or just the
positive region when
determining relative fluorescence intensity values.

Any advice would be appreciated.

Thank you,

Stevan Lauriault, B.Sc.
Research Assistant
Trauma Group
Operational Medicine Section
Defence R&D Canada - Toronto
Stevan.lauriault@drdc-rddc.gc.ca
Work:  416-635-2000 ext. 2303
Cell:  416-949-4066

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Received on Tue Oct 10 11:18:00 2006
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