FlowJo version 10 has a new preference setting that allows users to customize the data scaling based on the cytometer used to collect each file.
(*Please note – Instrumentation companies are constantly releasing updates to their software and do not always notify us of changes they have made to scaling keywords. If you cannot get your data to look like it does on the machine – PLEASE contact us right away)
Cytometers output FCS (Flow Cytometic Standard) files that contain keywords, information about the data organized by three letter codes. Keywords that are standard to all FCS files are prefaced by a $. All FCS files have the keyword $CYT which lists the cytometer used to collect the data. They also have the $SYS keyword which lists the software system that was used. Through these keywords FlowJo v10 has the ability to discern when acquisition software was used to create each data file, and then apply scaling preferences on a file-by-file basis.
To access these settings, click the Preferences button, then from the preference menu click on the Cytometers icon which is outlined in red in the figure below.
Once you click this icon, the window shown in the figure below will appear. The red numbers have been superimposed on the window as an index.
A list of cytometers is on the left of the screen, near the red number 1. Clicking on each cytometer shows the display settings for any file identified as having been collected by that cytometer. The top choice is “Generic”, and will be used by default whenever FlowJo cannot determine which cytometer was used for acquisition.
Section 2 identifies the keyword values that FlowJo uses to determine which cytometer has been used. These cannot be modified.
Section 3 allows the user to select whether they would like the scatter parameters to be displayed by default on a linear or logarithmic scale, what the range of all parameters shall be, and whether the data should be scaled. The values recommended by the individual cytometer manufacturers have been implemented here by default, and so in most cases the user will not need to adjust these values. The “Divide by…” choice has been included to accommodate users of cytometers that scale their data down for display in the acquisition software, but record the actual measurements, such as most Beckman-Coulter products. By using a scaling factor here the data will appear the same as when acquired. Anyone who would like to see the actual recorded values can simply set this to 1.
Section 4 remains grayed out until the “Enable Transforms” check box is highlighted. Once checked, the user can specify the default number of positive and additional negative decades, as well as the width basis. For help on what each of these settings do, see the transform overview page.
For descriptions of each modern acquisition software and how to manually adjust the data to match the acquisition software’s display in FlowJo, click the appropriate link below: