Ask: I want to share the results of my experiment to confirm the transfection rate of hek 293 cells (suspended not adherent) with a plasmid that reports with GFP, 2 days and 5 days after transfection, I have 2 problems:1- cells changing their scattering from day to day (I am not sure if my gating was right or not). 2- The cells gave high fluorescence 4 days after transfection, it worth to say, when I directly ran flow without processing of cells (without wash, pbs or even spindown), the autofluorescence disappeared, is it something related to the dead cells.