RE: "Fl1 drift over time"

Hi Katrina,

I would be interested in hearing your replies to this problem.

We also have an FC500 and have noticed a very high shift in some
samples. We tend to see FL2 (although we have also seen FL1 and once
FL4) shifted out at least a log to a log and a half. It does not happen
on our unstained sample tube (which we routinely run on every sample due
to this issue) and does not happen on the isotype control. Sometimes
when we let the sample sit and repeat it the problem disappears and
sometimes it does not. When it happens, it happens for every tube in
that patient's panel but not for every patient either that day or even
on that carousel. We have been in contact with Beckman coulter several
times and have not yet figured this problem out. It does seem to happen
more in the afternoon when the instrument has been running for quite a
long time. I know that Beckman will tell you that it is temperature
stable, but I think there is some effect.

Thanks,




<mailto:kimberly.pacella@healthnetworklabs.com> 

Kimberly A. Pacella, MT(ASCP)QCYM

Manager, Immunology & Flow Cytometry

Health Network Laboratories

ph: (610)402-5593

e-mail:kimberly.pacella@healthnetworklabs.com

________________________________

From: Katrina Ann Walsh [mailto:kawalsh@unimelb.edu.au] 
Sent: Tuesday, May 08, 2007 10:16 PM
To: cyto-inbox
Subject: "Fl1 drift over time"



Dear Flowers, 


We have encountered a perplexing problem with our flow cytometer, a
FC500. 

We have noticed that the fluorescence intensity of back ground controls,
such as isotype controls or unlabelled cells appear to increase rapidly
over time, in the order of doubling every 20 seconds. This is quite
troubling when setting negative background, especially when positive
staining is a shoulder rather than a distinct population.

We notice that it occurs on fl1 and subsequently fl2, however it is not
obvious on fl4. We are told our 488 laser is behaving correctly.

We have tested epithelial cell line, lymphocytes and cyto-comp (Beckman
Coulter) lymphocytes and find large cells, but NOT small beads, exhibit
this "fl1 drift" over time. 

We have also tested buffer ph and resuspending cells in isoton. We find
that MFI fluctuates when the same samples are re-read and that this
problem occurs most often in the evening, but not always. 

Often we do not seem to have a problem at all.	The flow cytometer room
is air-conditioned  reasonably well and would not get over 27 during the
summer.

What could be the problem? All suggestions welcome. 


Thank you 


Katrina Walsh

School of Dental Science

University of Melbourne. 

email: kawalsh@unimelb.edu.au


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