RE: Hoechst vs FITC: possible?
kukuruga%kasle1.dnet.wayne.edu@rocdec.roc.wayne.edu
Thu, 8 Feb 1996 09:59:02 -0500
H33342 with FITC is only possible, I believe, with a"time" displaced
signal. If you run a fluorescence spectrum on H33342, you will find a
relatively complex emmission profile, with significant fluorecence
at 530nm. This is also more problematic due to the variabliity of
absorption/emmission with time of staining and concentration of the
dye. If you need to acquire these parameters colinearily, use a
Red-shifted dye, like 7-AAD.
I routinely use H33342 at approx. 10 uM. I have tried to go as low
as 5 uM and lost stoichiometry. This was in viable L1210 cells --
these cells were sensitive to H33342 at 10 um, but 5 uM was
inadequate for labeling. For fixed cells, it's important to note
that dye accessibility is improved to the extent of requiring at
least a 10x dilution of the H33342 -- perhaps more (needs to be tested).
CD ROM Vol 2 was produced by staff at the Purdue University Cytometry Laboratories
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If you have any comments please direct them to Dr. J. Paul Robinson, Professor & Director, PUCL,
Purdue University, West Lafayette, IN 47907. Phone:(317) 494-0757; FAX (317) 494-0517; Web http://www.cyto.purdue.edu
EMAIL robinson@flowcyt.cyto.purdue.edu
