Coulter XL
Rehse, Mark (REHSEMA@cellpro.cellpro.com)
15 Feb 95 09:30:38 PST
This message is in response to the inquiry by Brian Smith regarding
fluorescence compensation on the Coulter XL. We purchased as XL 18 months
ago. We optimize instrument voltages and fluorescence compensation on a
monthly basis using buffy coat labeled with antibodies against
numerically abundant epitopes such as CD3, CD4, CD19. We use antibodies
directly conjugated to PE, FITC and RED613 and we throw in PI as a fourth
color. On a daily basis we check instrument performance with beads to
make sure they fall in the target channels established with the labeled
buffy coat cells.
Using this optimization protocol I find that the instrument is correctly
setup for 90% of our applications. Users do not have to change voltage or
compensation values to run their blood or bone marrow samples. The
remaining 10% run samples which require changes due to cell type, etc.
On occasion I find that the PE fluorescence is slightly over or
undercompensated using the optimal settings. However, rather than it
being and instrument related problem, we have found this to be due to
differences in reagent quality and staining protocol. Different antibody
conjugations, different F/P ratios, number of washes, and fixation all
can lead to different levels of background staining and fluorescence
intensity. We find the XL instrument performance to be of high quality
both in terms of precision and accuracy as well as "user friendliness".
Hope this helps.
Mark Rehse
Rehse@CellPro.CellPro.com
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