I have a basic data analysis question. I am measuring oxidative burst from PMNs
and Monos using DHR123. I am stimulating my cells with TNFa and treating them
with actives to inhibit ROS. My controls include DHR only cells and DHR/TNFa
cells. I have previously been comparing my compound treated cells to the
DHR/TNFa cells without subtracting out the baseline DHR stained cells. This is
how I've seen it reported in the literature, and I've previously been advised
not to do so. However, I have now been asked to subtract out the baseline DHR
stained fluorescence to compare the treated vs. untreated cells. I am not
comfortable doing this, but I'm not sure why I'm uncomfortable. So, here are my
questions:
1) Is it okay to subtract out the baseline fluorescence value from the
stimulated cells in order to analyze the data?
2) If not, please give me a good explanation to pass on to my supervisor
(non-flowist) so that I can get over this particular hurdle.
3) If so, please give me a good explanation so that I can deal with my
discomfort.
Thank you in advance....Jodi L. McKenzie-Kroeger
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