Fw: "Fl1 drift over time"

Hi Katrina,

we had quite a drift problem on our FC500. We have the ability to read 96 
well plates and if reading in rows, could see a marked shift in signal 
between the columns. In our case it was due to temperature. We now have an 
additional fan fitted to the back of the machine to cool the machine down. 
Since then we have not seen the problem anymore. Could this be the same in 
your case?

regards,

Marie-Ange Watson
GlaxoSmithKline
Stevenage
United Kingdom





----- Forwarded by Marie-Ange E Watson/PharmRD/GSK on 11/05/2007 08:01 
-----

"Kimberly A. Pacella" <kimberly.pacella@healthnetworklabs.com> 
09-May-2007 20:14
 
To
"Cytometry Mailing List" <cytometry@flowcyt.cyto.purdue.edu>
cc

Subject
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,
 
 
 

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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