Hello everyone.
I'm currently doing some lymphocyte proliferation assays with CFSE. I cultured 1X10^6 cells per well and stained with 5mM CFSE and stimulated them with certain Antigens including CONA acting as a positive control. I just wanted to ask how many events should I run on the flowcytometer to obtain clear histograms? Or is there was a way to run a high number of events (e.g.50,000) and see how the graph would be at like 10,000 or 15,000 events on softwares like Flowjo or FCS express?
For a proliferation assay I would always aquire at least 50.000 events, better more. There is really no advantage of having fewer cells aquired - and should you really desire to look at fewer cells during analysis, you can always select that option in FlowJo (either display a given number of cells on a dot plot, or use a downsample gate). Good luck with your assays!
I would honestly acquire everything in your tube. Rather have more than fewer. If your starting was a million cells, you should have plenty at the end.
You can choose to view less events in a plot in FCS Express by formatting the specific options of the plot: http://www.denovosoftware.com/site/manual/index.html?specific_options.htm . The statistics in FCS Express however will always be based on the full data set. Another other option is simply to smooth your histograms or data...however it is not recommended to smooth a proliferation plot in FCS Express, you want the proliferation fitting to be based on as many events as possible.
For standard flow cytometry data generation, you should acquire around 100000 cells / plot.
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