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IT'S GOTTA BE THE SHOES!!!!!
He couldn't remember learning anything about shoes at his cytometry
training course, but, there were several other messages that had suggestions
for things to try...
1) "Do you know the way to San Jose?"
Having the good fortune of having a cytometer available from the "other"
company downstairs, he stained some cells with the antibodies in question,
and ran them, only to find that the tandems were dim to the same extent as on
his "Hialeah HotRod". BTW - Larry does not work in San Jose.
2) "Do you have this in another color?"
He received several messages extolling the virtues of PE-Cy5, and a kind
soul explained to him that tandem conjugates work by resonance energy
transfer, not by fluorescence emission. He bought the PE conjugates of the
antibodies in question, and found that they stained a lot more brightly than
did the PE-Cy5. He triumphantly faxed the histograms to Larry, who in turn
triumphantly faxed back *his* histograms, showing that PE-Cy5 stained with
the same intensity, if not brighter, as did PE. Are you detecting a trend?
3) "Get 'em while they're hot?"
He received several messages suggesting that conventional PMT's had low
qunatum efficiencies in the red, and that he could order a special PMT
(gallium arsenide tube) to more efficiently measure this signal. He figured
that was a little more than he should have to do. Someone else suggested that
he switch his filter blocks around to try and pick up the signal in a
"downstream" PMT. He did that, but it didn't improve anything. He called
Moe and asked him to send him *two* new PMT's (having traveled this route
before he knew *one* wasn't going to be enough..."wise guy - eh?"). One of
the PMT's was the wrong one, even though it had the "right" part number. The
other one still didn't improve the signal, but curiously required 100 volts
less to obtain the same signal intensity.
4) "Through a Glass DARKLY?"
Several others suggested that perhaps Larry was a bit remiss in suggesting
the use of a 645 LP absorbance filter. Also, that perhaps the dichroics in
the light path were suspect. So he called Moe and got him to send him a new
set of filters, and, to order a higher wavelenth filter for the PE-Cy5
detector, only to find that the longest longpass Moe had was
630..."Rats"...So he called an optics company just down the road, and ordered
a 670 bandpass filter, and tried that, but the signal was not increased
compared to the 645 LP.
5) Can you say "crosspost"?
Poor Curely failed to realize that he had sent his original query out while
he was reading newsgroups - rec.sports.basketball to be exact. He finally
figured *that* out, but just to be on the safe side, he went out and bought a
pair of Air Max's, because he surmised that maybe these shoe guys were on to
something, and, he knew that the *other* bald guy wears them, and, when he
heard him say, "I was going to sue her for defamation of character, but then
I realized I have no character"....he thought - Hey! *I* coulda said that.
The moral of the story?
a) It's probably not realistic to expect that your cytometer vendor will give
you the time of day if you're using reagents from another vendor.....after
all, the money's in the "juice".
b) It's probably equally unrealistic to expect that you'd be able to buy
*al*l your reagents from *one* vendor. Some just make some things better
than others, and vice versa.
Sorry to take up so much bandwidth, and sincere thanks for all the extremely
helpful suggestions.
--marc
Marc Langweiler 603-650-4924
DMS Pathology Lebanon, NH
langweiler@dartmouth.edu
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