We are performing cell cycle analysis using DAPI staining with flow cytometry. Although the cell density, preparation and dye concentrations are kept identical for all groups, we observe that the G1 peaks appear at different positions in the histograms across the different cell groups. Due to this variability in peak position, we find ourselves needing to adjust the analysis conditions for each sample individually.

Could this be due to technical factors related to flow cytometer settings or potential sample preparation issues that are not apparent? We are considering adjusting the conditions applied for analysis, but I am unsure if this is the appropriate approach. What could be the underlying causes of such variability, and are there recommendations for standardizing conditions to ensure more consistent G1 peak positioning? Any insights or suggestions for standardizing the analysis would be greatly appreciated.