I'm working with HEL and CMK (megakaryocytic cell lines) which 'naturally' become polyploid. I am currently using Hoechst 33342 to stain and am running the samples through a Beckman Coulter Quanta SC.
My current problem is doublet discrimination. I am not sure how to do it on the Quanta SC. The conventional way would be using fluorescence area vs width. However the problem is that the Quanta SC does not save information on fluorescence width so I am unable to use this method.
I do know that there are doublets, as I see a small peak between my 4N and 8N population on the histograms. I've tried gating on side scatter vs fluorescence concentration, but the 'doublets' are sandwiched right between the populations and are almost impossible to distinguish.
I've tried asking technical support for help, but so far they have not come up with any satisfactory solutions.
Doublet Discrimation on the Beckman Coulter Quanta SC
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