From: sperfetto@hiv.hjf.org
Date: Fri Mar 10 1995 - 09:11:29 EST
Typically, our 2nd laser (green HeNe) has CV's very close to the 488nm beam ie.
<1.0, this of course does not include the FS detector since our beams are not
co-linear. The HeCAD laser usually has higher CV's around 2.5, but for
calcium work we find this adequate.
Beads used; Coulter DNA check
Instrument : ELITE-ESP / 3 lasers with gated amp.
SP
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Subject: 2nd laser CVs
Author: t-delohery@ski.mskcc.org (Thomas Delohery)
Date: 3/9/95 6:50 PM
Hi,
I'm conducting an informal survey to compare the differences
in CVs for fluorescence of either nuclei, Coulter's "DNA Checks"
or any other extremely uniform fluorescent particle between 1st
and 2nd lasers (or 3rd?).
I'm assuming the CVs for the 2nd laser will always be higher than
the 1st on standard commercial instruments. (Say it ain't so.)
My assumption is primarily based on the fact that the 2nd beam
is not passing through the center of the laser focusing optic.
I'm currently using a FACStar Plus and can't seem to get the CVs
for the 2nd laser even close to those of my 1st. I'm currently in
the process of dismantling and realigning the entire system. So I'd
really be interested in hearing from anyone who routinely uses the
2nd laser for linear fluorescence measurements on ANY flow cytometer.
I just want an idea of what is the best I can expect to achieve.
For instance, I routinely obtain full CVs less than 2% on green,
orange and red fluorescence with Coulter's "DNA Checks" with the
1st laser. If the 2nd laser is tuned to 488 or 514nm (same power
and optical filters) what is the best I can hope for? CVs of 3%? 8%?
Before the dismantling I was looking at CVs around 15%!!!
Any response would be greatly appreciated.
tom d.
"At the heart of the slaughterhouse - always - enough room to nourish awe."
- J. Nichols
--
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Thomas Delohery | Internet: t-delohery@ski.mskcc.org
Manager, Flow Cytometry Core Facility | Phone: (212) 639-8729
Memorial Sloan-Kettering Cancer Center | Fax: (212) 794-4019
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