autofluorescence & isotype control fluorescence

From: Karim Vermaelen (Karim.Vermaelen@rug.ac.be)
Date: Fri Oct 20 2000 - 03:27:26 EST


Hi everybody,

Here's maybe a tricky one:

Suppose you got highly autofluorescent cells and the isotype control is
in the 2nd-3rd decade. The same isotype control in low autofluorescent
cells is under the 1st decade.
Now suppose the signal for marker X on these high autofluorescent cells
is also in 2nd-3rd decade, and exactly overlaps the isotype control.

What's the correct interpretation of these results?
1) there's no difference between isotype control and marker X, so the
cells are negative for marker X
or
2) marker X is in the 2nd-3rd decade, so it's definitely positive
regardless of isotype control background

Another way to state the question: are fluorochrome fluorescence (be it
Ag-specific or aspecific binding) and autofluorescence additive signals
for the same cell?

TX a lot for any clue!

Karim
Pulmonary Immunobiology
Ghent University Hosp.



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