I have numerous FCS files spread across multiple directories. I originally wanted one set of plots in the first row, and the same set of plots duplicated in the second row. I could then compare between directories by cycling through samples in each row.
Is it possible to copy regions/quadrants/markers from one plot to another? I know you can create a single plot with a region/quadrant/marker, then duplicate the plot, but if you want to change the position of the region/quadrant/marker in both plots afterwards, is that possible? I couldn't find any quantitive description of quadrants and markers. I could find a description of the region(s) in the "Edit Regions/Gates" menu, although this didn't seem editable.
Is the idea in FCSalyzer that we should have all our FCS files in a single directory? This can be a bit fiddly with many plots or multiple experiments.
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Quadrant and markers are used to identify relevant populations in a given plot. They work on a per-plot basis and are independent of each other. They are also independent on the parameters that are shown in the plot. They are used for statistical analysis for a given plot.
Regions identify relevant populations for the whole analysis/document. One region applied to all datafiles and all plots. If a region is changed, this change applies to all plots and datafiles. Regions are defined based on specified parameters. Regions are used for gates (see below) and for statistical analysis.
Which one to use depends on your needs:
If you want to have an identical region-of-interest for many plots, it might be best to create a region. However, there are no regions for histograms. If you want to be able to slightly adjust the region-of-interest in various plots (for example because some samples have more debris or such), then quadrants/markers are the better choice.
Keep in mind: regions are always applied to all datafiles and plots, even if they are not visible. The statistics dialog will always calculate them, if "regions" is selected. A change to the region will propagate to all plots. Quadrant and markers apply only to the plot they are visible in and they are not connected. So a change in one quadrant will not influence a quadrant in another plot.
Regions are also the building blocks for gates, which are used to filter events for analysis and visualization. The "Edit regions/gates" dialog is meant for that: to rename regions, to delete them and to create/delete/name gates.
It should be possible to copy markers, regions and quadrants between plots. This is done with the "Edit" menu, and there are also keyboard shortcuts. Please see here: https://sourceforge.net/p/fcsalyzer/wiki/Usage/ and scroll to the "Edit" section.
In short: select the marker/region/quadrant, then make copy (I am not sure which keyboard shortcut that is in Linux, under Windows it's CTRL-C), select the plot where you want to add the marker/quadrant and paste (CTRL-V). There can be only one quadrant per plot, so an existing quadrant will be replaced (updated). There can be multiple markers in one histogram, so existing markers will remain and the new marker will be added to the plot.
2) Files from multiple directories / experiments in one analysis
That should be possible. Just as you wrote, you can make a plot with a datafile from one directory, and a different plot with a datafile from a second directory, and then it should be possible to cycle through each directory in parallel. I wanted it to work like that, please let me know if there is a bug.
However, please kkep in mind that all datafiles should be of the same type, especially concerning range and resolution. If not, things can get difficult when using regions and gates. But most probably that is not an issue for most users, as users tend to have multiple experiments from the same machine - which will produce similar datafiles. Mix n Match from different cytometer models (BD vs Miltenyi vs Biorad vs ...) is not a good idea.
If you would like to refer to this comment somewhere else in this project, copy and paste the following link:
Sorry for the overload with the questions! And thank you again for the prompt and informative reply. That answers all my questions perfectly. Thank you!
If you would like to refer to this comment somewhere else in this project, copy and paste the following link:
I have numerous FCS files spread across multiple directories. I originally wanted one set of plots in the first row, and the same set of plots duplicated in the second row. I could then compare between directories by cycling through samples in each row.
Is it possible to copy regions/quadrants/markers from one plot to another? I know you can create a single plot with a region/quadrant/marker, then duplicate the plot, but if you want to change the position of the region/quadrant/marker in both plots afterwards, is that possible? I couldn't find any quantitive description of quadrants and markers. I could find a description of the region(s) in the "Edit Regions/Gates" menu, although this didn't seem editable.
Is the idea in FCSalyzer that we should have all our FCS files in a single directory? This can be a bit fiddly with many plots or multiple experiments.
Many questions in one post :-)
1) Regions, quadrants, histograms
Quadrant and markers are used to identify relevant populations in a given plot. They work on a per-plot basis and are independent of each other. They are also independent on the parameters that are shown in the plot. They are used for statistical analysis for a given plot.
Regions identify relevant populations for the whole analysis/document. One region applied to all datafiles and all plots. If a region is changed, this change applies to all plots and datafiles. Regions are defined based on specified parameters. Regions are used for gates (see below) and for statistical analysis.
Which one to use depends on your needs:
If you want to have an identical region-of-interest for many plots, it might be best to create a region. However, there are no regions for histograms. If you want to be able to slightly adjust the region-of-interest in various plots (for example because some samples have more debris or such), then quadrants/markers are the better choice.
Keep in mind: regions are always applied to all datafiles and plots, even if they are not visible. The statistics dialog will always calculate them, if "regions" is selected. A change to the region will propagate to all plots. Quadrant and markers apply only to the plot they are visible in and they are not connected. So a change in one quadrant will not influence a quadrant in another plot.
Regions are also the building blocks for gates, which are used to filter events for analysis and visualization. The "Edit regions/gates" dialog is meant for that: to rename regions, to delete them and to create/delete/name gates.
It should be possible to copy markers, regions and quadrants between plots. This is done with the "Edit" menu, and there are also keyboard shortcuts. Please see here: https://sourceforge.net/p/fcsalyzer/wiki/Usage/ and scroll to the "Edit" section.
In short: select the marker/region/quadrant, then make copy (I am not sure which keyboard shortcut that is in Linux, under Windows it's CTRL-C), select the plot where you want to add the marker/quadrant and paste (CTRL-V). There can be only one quadrant per plot, so an existing quadrant will be replaced (updated). There can be multiple markers in one histogram, so existing markers will remain and the new marker will be added to the plot.
2) Files from multiple directories / experiments in one analysis
That should be possible. Just as you wrote, you can make a plot with a datafile from one directory, and a different plot with a datafile from a second directory, and then it should be possible to cycle through each directory in parallel. I wanted it to work like that, please let me know if there is a bug.
However, please kkep in mind that all datafiles should be of the same type, especially concerning range and resolution. If not, things can get difficult when using regions and gates. But most probably that is not an issue for most users, as users tend to have multiple experiments from the same machine - which will produce similar datafiles. Mix n Match from different cytometer models (BD vs Miltenyi vs Biorad vs ...) is not a good idea.
Sorry for the overload with the questions! And thank you again for the prompt and informative reply. That answers all my questions perfectly. Thank you!