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Tom Mchugh (mchugh@pangloss.ucsf.EDU)
Wed, 12 Apr 1995 15:12:01 -0700 (PDT)

I am coupling FITC, PE and PE-Cy5 to beads of various sizes. I would like to
use these as standards of known number of fluorochromes per bead. I am
looking for a method to approximate the number of fluorochromes per
bead. Is it reasonable to run the soluble fluorochrome, trigger on
fluorescence and plot the number of soluble fluorochromes versus the
mean of the fluorescence histogram. And then use the mean fluorescence
associated with a known number of soluble fluorochomes
when I run the beads as a "mean equivalent soluble fluorochome" in order
to assign an approximate # of fluorochromes to each bead?
I have run the Flow Cytometry Standards beads along with mine but do not
want to buy theirs for the number of studies that we would like to do
and the range of fluorescence they offer is too narrow.
(I do acknowledge the difficulty of determining antigen concentration
per cell due to the antibody affinity, number of binding sites as well
as the fluorochrome:protein ratio).

Thanks,

Tom McHugh
Dept. Lab. Medicine
Univ. Calif., San Francisco
mchugh@labmed.ucsf.edu

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