When making multiple matrices like this, you only need to repeat the drags of the successive Cy5PE populations followed by steps 4 and 5, saving a different matrix for each set of reagents. You will need to collect a singly-stained compensation control sample for each different Cy5PE reagent, and then construct a different matrix for each of these. Different Cy5PE reagents (even from the same manufacturer) have different spectra and thus require different compensations. This is true, for instance, when you are using different lots of tandem dyes (like Cy5PE). A FlowJo workspace lists all of the sample data files that you load, displays the gates, statistics, and other analyses that you apply to those samples, and provides access to tools, such as graphical layouts and statistics tables, that you can. The compensation matrix is represented by the grid to the left of the sample name. FlowJo Software is an integrated environment for viewing and analyzing flow cytometric data. In some cases, you will need multiple compensation matrices for the same experiment. Back to product Applying a compensation matrix Create new, apply old, or compare different matrices. You can follow this example on your own computer by downloading the Compensation Demo Data. FlowJo Software The FACSVantage SE was configured as follows. This includes understanding embedding and using keywords, the FlowJo compensation wizard, spillover spreading matrix, FlowJo and R, and creating tables in FlowJo. compensation controls, use FACS staining buffer alone, or with a 1 100. Display the compensated parameter (in brackets ) on the graph axes. FlowJo is a powerful tool for performing and analyzing flow cytometry experiments, if you know how to use it to the fullest.Apply the compensation matrix to the appropriate samples.Drag each of the positive and negative populations from the workspace window into the appropriate box in the compensation dialog window. Chapter 6: Unmixing and Compensation 59 Negative/Unstained Controls In addition to positive reference controls needed for spectral unmixing, an unstained control is also necessary to assess autofluorescence.Bring up the Compensation Definition dialog window, or Compensation Wizard.Set a gate for the positive and negative populations for each singly-stained sample.This is a simple example showing how to do three-color compensation in FlowJo. This tool works for matrices created in FlowJo, as the single stain controls are needed to calculate several statistics from to produce each value in the SSM.
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