[Cytometry] Consistent difference between CST vs Manual laser delays

Ian Dimmick ian.dimmick at newcastle.ac.uk
Wed Feb 19 09:28:29 EST 2014


Hello Chuan, firstly I think if you check laser delay settings you should probably use the CS&T beads if they are what your laser delays have been set up on initially. 

I then would perhaps check the laser delays , and expect some minor drift , in fluorescence during the day due to temperature changes , but this will hardly ever affect the analysis of your cells, only in the scenario perhaps where you have applied a very narrow signal window extension, thus seeing a difference which is more proportional to bead/ cell size rather than anything to do with laser/ instrument drift 

Good luck 

Ian 


>-----Original Message-----
>From: cytometry-bounces at lists.purdue.edu [mailto:cytometry-
>bounces at lists.purdue.edu] On Behalf Of Chuan En Lam
>Sent: 19 February 2014 00:22
>To: cytometry at lists.purdue.edu
>Subject: [Cytometry] Consistent difference between CST vs Manual laser
>delays
>
>Hi all,
>Recently we seem to be seeing consistent differences between CST set laser
>delay values vs Manual laser delay values on our AriaIIu.
>
>Instrument setup/details:
>AriaIIu (Blue/Red/Violet laser) inside ClassI CAS hood.
>Morning cold temp: 20.0deg,
>Start up to CST: 22.0 deg
>2hrs after start up to end of day: about 24.0 deg
>
>Mostly, after initial fluidics startup, CST would be run. We then double
>check with manual laser delay setting using single peak beads and 8 peak
>rainbow beads.
>
>We found a consistent requirement to shift the laser delays by about 0.5
>compared to CST to get optimum position. This is in general consistent
>across nozzles (70um, 100um). We are not sure if temp changes has an
>effect, or was the flow cell dirty (have done flow cell cleans with
>Decon/Coulter cleanz).
>
>In one scenario, 70um nozzle, after flow cell clean.
>
>_________CST AM____Manual
>AM____12PM____4PM____4pmCST____4pmManual
>Red           -43.06            -43.50           -43.50      -43.50
>-42.86            -43.36
>Violet          40.64             40.18            39.92
>39.92         40.46             39.86
>
>
>
>
>Has others seen similar effects or ideas what this might indicate?
>
>Normal condition sorting (log scales etc) seems to be ok with clean
>purities.
>
>Regards,
>Eric
>
>
>
>
>Eric Lam
>Flow Cytometry Lab Technician
>Garvan Institute of Medical Research
>Sydney, Australia
>_________________________
>http://www.flow.garvan.org.au
>Ph 9295 8431
>e.lam at garvan.org.au <r.salomon at garvan.org.au>
>_________________________



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