Re: best multicolor instrument?

From: Andy Riddell (ar289@hermes.cam.ac.uk)
Date: Thu Oct 12 2000 - 05:36:51 EST


>	I would like to get the group's opinion on the best available
>instrument for doing multicolor [ at least 6, in the near future maybe
>7, and potentially 8 ] analysis as to
>Altra vs Vantage vs Moflo
>
>1] how well does the 8x8 compesation matrix on the Moflow work?
>
>2] if I need to use aftermarket software compensation , is there any
>advantage to Mac based data vs. Windows based data?
>
>3] is there a big difference in sensitivity between the jet in air flow
>cell [Moflo &BD] and the quartz [BD & Coulter] for reagents such as
>Cascade Blue and APC-CY7?
>
>4] which has the best design for multilaser alignment?
>
>5] which has the best disign and optics for signal collection?
>
>Thanks for any info.
>
>Regards,
>Tom
>
>--
>*****************************************************************************
>Thomas W. Mc Closkey, Ph. D.
>Director of Flow Cytometry, North Shore University Hospital
>Assistant Professor of Pediatrics, New York University School of
>Medicine
>Boas Marks Biomedical Research Center, 350 Community Drive
>Manhasset, Long Island, New York 11030
>ph:  516-562-4844 [office], 516-562-1135/4641 [lab]  fax:  516-562-2866
>*****************************************************************************

Hi Tom,

I'll try to answer your questions one at a time where I can.


1] how well does the 8x8 compensation matrix on the MoFlo work?

-- The compensation matrix is quite good on the MoFlo, there is a little
distortion of the populations due to the algorithm, but it is really of no
significance. I built analogue compensation circuits for my MoFlo couple of
years ago because;
a) The software compensation algorithm on the MoFlo was dreadful. It has
much improved and they have moved the compensation matrix onto a DSP chip
that now allows you to sort on compensated parameters.
b) Then, you couldn't sort on compensated parameters.
Although my analogue electronics worked well it did have a few problems
with phase alignment of pulses which was exacerbated at high signal input
into the compensation. And it was not capable of interbeam compensation.
Cytomation, in utilizing a DSP chip and improving their algorithm, have
sorted their problems out. The DSP has a number of advantages over the
analogue solution.  The DSP does complete interbeam compensation. DSP
outputs are saved along with the uncompensated data allowing you to go back
after acquiring your data and re-adjust the compensation. The overhead is
that your file size is doubled, but that is no real pain when you have 12GB
at your disposal. The DSP has no pulse phase problem.

2] if I need to use aftermarket software compensation , is there any
advantage to Mac based data vs. Windows based data?

-- I'm keeping out of this one... I like Acorn RISC OS, but that's another
thread!

3] is there a big difference in sensitivity between the jet in air flow
cell [MoFlo &BD] and the quartz [BD & Coulter] for reagents such as ascade
Blue and APC-CY7?

--I haven't used these reagents so I cannot comment. However, I ran some
really ancient 5-peak calibration beads to compare sensitivities of my
MoFlo and my FACSCalibur (running at 35 psi with 100u nossle) I've enclosed
the results. The MoFlo was better but I do think that my BD machine needs
realigned.

4] which has the best design for multilaser alignment?

-- In my mind MoFlo for high speed sorting, because each beam is
independently steered and focused. I can't comment on the Altra as I have
never seen one running, but I think they use independent beam steering and
focusing (please correct me if I'm wrong).

5] which has the best design and optics for signal collection?

-- Again I would say the MoFlo, because the optical bench, electronic rack
and the collection optics are open so you can easily add more PMTs, ADCs,
etc,.

I hope this helps.

Best Regards,

Andy.


Andy Riddell Flow and Imaging Lab CIMR Hills Road Cambridge UK tel: (0)1223 762597 fax: (0)1223 336900 email: ar289@cam.ac.uk



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