RE: FACS question

From: Mark Kukuruga (kukuru@med.umich.edu)
Date: Tue Dec 18 2001 - 11:01:00 EST


. . . the data always gets prettier at closing time . . .

But seriously, what we're talking about here is holding ourselves to a higher standard -
- and what's wrong with that?  We've had a month's worth of discussion over good/bad
publications, but nobody's looking at the cause . . . simply that there are confused
operators/users in the field - - and many don't even know they're confused.
Blame is not what we're about, but rather, what can we do to eliminate as much of the
confusion as possible?  Certainly training is key, courses are a plus, but ultimately we
need a commitment from those of us who operate instruments to understand our technology
to the maximum extent of our ability.
Back in the day, when everyone had an "open" bench, understanding came from hands-on
manipulation.  With the development of turnkey analyzers, we lost that basic connection
to the bench's parts, and with it the understanding of how those parts are important
to the process.  I've complained about the use of the arbitrary "FL" designations in
the past as well, not because the questions are trivial, but rather that it represents
a basic lack committment to understanding.  When someone asks a question about why
propidium iodide, when read in "FL2," bleeds into "FL3," it's obvious they don't really
know what FL2 and FL3 are.  If that's true, how can they be expected to use the flow
cytometer correctly?
Ultimately, I feel that part of the good/bad data problem lays somewhat with our
failure to be harsh when it's warranted.  In general, one rises to the level of the
competition . . . and greater demand calls for greater effort.
MAK.

--
Mark A. KuKuruga, Managing Director
University of Michigan Flow Core
7416 CCGC 0946
(734) 647-3216, fax (734) 936-7376
kukuru@umich.edu


>>> Andrew Beernink <ABeernink@novasite.com> 12/14/01 04:15PM >>>

True.  But can anybody explain why I get better results after consuming beer
Brand X or martini Brand Y?

-----Original Message-----
From: Mark Kukuruga [mailto:kukuru@med.umich.edu]
Sent: Thursday, December 13, 2001 8:00 AM
To: cyto-inbox
Subject: Re: FACS question



Paul . . . thanks for this response.  I've tried this, and it works well.
I've had some trouble finding a good source for reagent B though.
MAK.

--
Mark A. KuKuruga, Managing Director
University of Michigan Flow Core
7416 CCGC 0946
(734) 647-3216, fax (734) 936-7376
kukuru@umich.edu


>>> "J.Paul Robinson" <jpr@flowcyt.cyto.purdue.edu> 12/11/01 09:20PM >>>

Colleagues: I am going to answer this question in a manner that I hope you
all
understand (or not)......(think of the bad data issue)

You can solve the problem by adding reagent A to reagent B, plotting FL3 Vs
FL2.
If you add reagent C, then a single histogram of FL5 should do the trick. I
think you
could also try probe A and probe B, both of which should have the right
spectra...Oh,
turn the power of laser 1 up a bit, and you should get some valuable
data.....

sorry, I couldn't resist it...
please stop using terms which are totally undefined....

Paul Robinson


On 10 Dec 2001, at 11:58, Laura H
odges wrote:


I'd like to post a question on your website:


Can anyone suggest another P-gp accumulation probe
other than rhodamine 123 which has too broad of an
emission spectrum for my use.  I am trying to find a
P-gp probe that can be used with other fluorescent
markers in channels FL2, FL3, and FL4 in a multicolor
assay.


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