Compensation Tool

The Compensation tool is composed of three main parts: the Matrix Manager, the Sample Manager, and the Compensation Viewer.

Matrix Manager:

This is the pane in the upper left corner labeled Compensation Matrices.  This window allows you to create (+), duplicate (=) and delete (-) compensation matrices much as you would layouts or tables. You can also adjust the color label of the compensation matricies by clicking on their colored square and selecting a desired color.

Sample Manager:

This Sample Manager is directly below the Matrix Manager pane, and is labeled Samples.  This window allows you to assign different samples from your experiment to the various compensation matricies that you may create.  Once a sample has been dragged to the sample manager manager, FlowJo will assign it to the selected matrix in the Matrix Manager, and draw a colored compensation indicator line next to the sample name in the workspace, in the same color as the selected matrix. Samples can be deleted from this list by selecting them and pressing the delete key on your keyboard.

Compensation Viewer

The compensation viewer contains two tabs: Matrices and Transforms, and Wizard Graphs.  The first tab is customizable, allowing users to view particular aspects of compensation are most important to them. The different configurable GUI components for the
compensation editor can be accessed though the View menu in the compensation editor window. These components include the Show Samples/Gate Assignments, the Show Comp Matrix,  Show Comp Script and Show Transformation Settings, explained below.  In addition to the customizable components, the compensation viewer tab has two buttons, Apply and Finish. The Apply button propagates any changes that have been made to a matrix to any samples that have been assigned to that matrix. The Finish button finishes the process of creating a compensation matrix and divorces the link between the matrix and the compensation controls so that the matrix is not inadvertently altered by changes made to the compensation controls and their gates.

Sample/Gate Assignments:
This GUI component allows you to verify which samples and gates are being assigned to each role in the process of creating a compensation matrix. This is available from the View menu. Right clicking on this box allows you to adjust sample and gate assignments such that each population is being used for the correct role in your matrix calculation.  Adjusting the sample for each color will cause a change in both the gates and compensation matrix downstream.

Compensation Matrix:
This is the actual compensation matrix that FlowJo will use to compensate any samples assigned to it. This can be made visible from the View menu. In the majority of cases, you will not need to adjust these numbers by hand for any reason.  Do so with skill (statistics) and at your own risk.  To find a particular spillover value, look at the rows of the matrix, and match the stain that you want to adjust in other parameters by looking over at the list of Fluorophores to the left.  Traverse right across the row for this stain until you find the channel for which you would like to adjust the first fluor's contribution to.  This number is represented as a decimal, usually (but not always) between 0 and 1.  Small negative values are typically harmless.  Values greater than 1 may indicate incorrect samples assignment, or that your stain is simply brighter in a secondary channel than in the one that you are attempting to measure it in.  To adjust a spillover value in a flowjo compensation matrix, simply change the text in the desired field in the matrix to the desired value, and press apply.

Compensation Scripting
This tool is used for calculating compensation matricies automatically in a template by using keywords to identify your controls. This can be made visible by turning it on from the View menu.

Transformation Settings
This is a detailed view of the various biexponential transformation setting that will be used to optimize the appearance of your data.  We typically recommend using the transform button on the graph window rather than adjusting these values. This can be made visible by turning it on from the View menu.

The second tab of the compensation viewer is shows the wizard graphs to confirm that flowjo is making good gates based on your compensation controls.  Any gate that needs to be adjusted can be opened up in a graph window by double clicking on it.  Any changes made to a gate will propagate automatically throughout your workspace.