Increasing Cytoplasmic fluorescence

Walter Sharp 102675.320 at CompuServe.COM
Tue Nov 5 08:10:26 EST 1996

Dear All,
	Got an unusual problem that we see quite often these days.

When running cytoplasmic markers (TdT, Mu etc) we see the fluorescence intensity
of both the FITC stained cells and non specifically stained controls creep up
the 4 decade log scale, levelling out after 2 minutes and up by 1 decade.
Re-running the SAME tube shows the same pattern, starting at baseline and
levelling off 1 decade up.
No changes in FS, SS or aquisition rate are evident.
We never see this with surface staining at all and it doesn't happen with all
our Leuks.
The spectral spillover into FL2 (rpe) also shows a concomittant increase.
We have a four PMT Coulter XL with System II software.
We currently use Caltag's "Fix and Perm" with Mu and HT6 TdT clone from Dako.
We did see it once with our previous "in house" permeablisation method but it
never repeated itself.

Any ideas ?

Wal Sharp

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