Hi!
I´m quite new with FACS measurements; my question is: when I adjust the voltage of FSC, SSC and my fluorochrome, they need to be all different?
And for the fluorochrome there is a specific voltage?
Thank you!
Hi Stefania:
Yes, it is very rare the voltage for different parameter is the same.
To get the optimized voltage for each PMT detector:
1) CS &T beads to define baseline and performance of each detector;
2) Optimize each PMT voltage with commercial calibration beads (above two should be done by facility manager)
3) For you, negative control, positive control samples are important for you to choose your parameters;
4) Find some colleague or friend really know FACS around you to help you setup the experimental model, then your life will be easier.
Regards
My suggestions are
Since you seem to be a novice to use the flow cytometry , start with an help from experienced person. because if you do not adjust your PMTs properly there would be a difficulty in the compensation to pass. Moreover improper compensations can overrate or underrate the cell populations. if you have any other specific queries I would be interested to answer. I have worked with 13 parameter flow on human blood and biopsies for various immunological cell subsets. Good luck!!
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