[Cytometry] APC problems

Derek Davies derek.davies at cancer.org.uk
Fri Feb 27 14:59:09 EST 2009


Hi Chris,

What flavo(u)r of red laser do you have? If its a 648nm you may be picking
up direct laser light which would sort of explain why you get better
separation by moving the APC filter up a tad. Don't forget that bandpass
filters will let light in from outside their range - it may not be much but
if its high power laser it will reduce your sensitivity. We have had to be
very careful with optical filter selection on both my LSRs.

Derek



On 26/2/09 12:27, "Christian Willberg" <Chris.Willberg at ndm.ox.ac.uk> wrote:

> Dear All,
> 
> I was hoping somebody could help us with a problem we're having with
> our APC PMT on a new LSRII, our local BD engineer is at a loss.
> We have been struggling to get good APC stains with antibodieswe know
> work very well on other machines. When we checked with BD rainbow
> beads we noticed that the APC dim beads where bunching up halfway
> along the axis - this wasn't seen on either the Alexa700 PMT or APC-
> Cy7 PMT (please see attached PDF). We checked that this wasn't due to
> the filter (660/20) using a second 660/20 filter and the same pattern
> was seen. However, when we swapped the 660/20 filter for a 670/14 -
> the problem was resolved. Ideally we would like to use the 660/20
> filters for APC and not the 670/14. Can anyone explain what could be
> causing this and how we can fix it?
> 
> Many thanks,
> Chris Willberg
> 
> Chris Willberg
> Biomedical Research Centre - Immunology Theme
> University of Oxford
> Oxford, UK
> 
> Tele: 01865 271290
> or tele: 01865 222352
> e-mail: Chris.Willberg at ndm.ox.ac.uk
> ----------------------------------------------------------
> 
> 
> 
> 
> 
> 
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