Dual laser compensation

Thomas Knapp tknapp at scripps.edu
Mon Mar 11 14:09:43 EST 1996


Joseph Webster writes...


>Hi All,
>Has anyone found, made, thought of, a way to compensate between lasers
>on a machine such as a FACStar plus?
>We see bright PE appearing in the Texas Red channel, apparently excited
>by the dye laser.
>
>    Joseph.
>Joseph Webster (O.I.C. Flow Cytometry)                      ===
>Centenary Institute of cancer Medicine & Cell Biology


Before attempting compensation across time on a Facstar Plus, (a feature which
is obviously not built in to the instrument), I would consider the following 
possibilities:

(You probably already know these things, but for the benefit of those
who may have similar problems and questions...)

1. Is the dye laser tuned to a wavelength low enough to excite the PE? If so,
it might help to tune it to a higher wavelength to get the PE signal 
(excited by the dye laser) out of the TxRed channel.

2. Have you tried a different source of PE? It is possible that there is some APC
contaminating the PE. Both of theses fluorochromes are phycobiliproteins and may
have some cross contamination depending on the efficiency of their
separation by the manufacturer of the reagent.

3. Try to optimize the time base of the separation of the lasers and the positioning
and aperature size of the iris in front of the FL3-2 and FL4 detectors.



Tom Knapp
Flow Cytometry Core Facility
The Scripps Research Institute
tknapp at facs.scripps.edu
www http://facs.scripps.edu/

phone
(619) 554-8396,554-8286



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