Since 1994, Flow Cytometrists have been using FlowJo to analyze data collected on BD cytometers to create statistics and figures pivotal to publications on immune subsets. Until now, FCS files were the only data exported from FACSDiva™ and read into FlowJo – Now within FlowJo v10.6, FACSDiva™ experiments can now be imported into FlowJo and FlowJo can export FACSDiva™ templates.

This means that gating information and population hierarchies don’t need to be lost in the handoff. Additionally, you can now create FACSDiva™ experiments in FlowJo for experiment reiterations and precise cell sorting gates.

 

Importing FACSDiva™ Experiments into FlowJo

Once you have finished collecting your flow cytometry data on a BD cytometer running FACSDiva™, you will need to export your experiment file (.XML) by either right-clicking on the experiment and selecting Export-> Experiment from the dialogue or by choosing Export-> Experiment from File on the FACSDiva™ ribbon. You will be given the option to export as a Directory or a Zip File. Choosing Directory will export a folder with your experiment XML; Choosing Zip File will export a Zip file that contains your experiment XML and your FCS files. Please choose Directory, as this will be more easily loaded into FlowJo.

 

With FlowJo v10.6 open, load your FCS files from the experiment by dropping the files into the FlowJo Workspace or from the Add Samples dialogue. Next, right-click on any of loaded samples and select “Import from FACSDiva™” to open the Import from FACSDiva™ dialogue.

Once the Import from FACSDiva™ dialogue has been opened, you will be asked to choose:

  • The experiment using the file navigator
  • The Global Worksheet with the gates that you are importing
  • An FCS file with the scaling and compensation that you would like applied
  • Whether you would like to import compensation and scaling only

 

After making these decisions, you will then need to click the “Import from FACSDiva™” button to bring the information into FlowJo.

Export from FlowJo to FACSDiva™

When you have created the gating structure in FlowJo v10.6+ needed for repeating your experiment, you can modify an existing Diva experiment (.XML file) that  and import that file into FACSDiva™. Simply right-click on a sample that contains the desired population hierarchy and gating, and from that dialogue, click on “Export to FACSDiva™.” A new window will open with the following options:

  • Create a new experiment or modify an existing experiment
  • Select the FACSDiva™ experiment to modify
  • Select the output directory for the new experiment
  • Select the gates to export
  • Choose the plot style foe the new plots
  • Apply compensation and scaling to all files in experiment
  • Only display the last population gate

 

Once you have made your selections, press the “Export to FACSDiva™” button to create the Folder containing the FACSDiva™ experiment XML.

 

Some Technical Details

  • Only the following transformations are supported: Lin, Log, and Biex. Make sure you’ve properly scaled parameters prior to gating in FlowJo, by accessing the Customize Axis dialog in the “T” button next to either axis in Graph Windows before gating.
  • Only the following gating types are supported for the conversion: Range, Bi-sector, Quad, Polygon, Rectangle, and Ellipses. Note: Ellipses converted into FACSDiva™ XML will turn into polygons to approximate the ellipsoid shape.
  • Parameter naming is important for the conversion of FlowJo’s gates into FACSDiva™ (and vice versa). In other words, the experiment’s FCS files need to match.
  • Statistics calculated on populations in FlowJo will differ from those calculated in FACSDiva™ for the same populations. This difference is not expected to vary outside biological significance.
  • Values in compensation matrices in FACSDiva™ are rounded to a few decimal places for display purposes only, but they are stored in full precision for calculating spillover subtraction.
  • Compensation matrices generated with spectral optimization (aka “overdetermined compensation”, or “non-square comp. matrices”) are not supported in FACSDiva™.

If you have any questions about how FlowJo can help facilitate a seamless workflow from acquisition to analysis and back again, please reach out to the Product Innovation team with your questions or ideas: flowjo@bd.com