How do I fix the biexponential axes on a plot?
The objective of visualizing data on a measured parameter is to evaluate the distribution of the sampled data. The axis scaling on a plot only affects how data is displayed on the plot with statistics always derived from the raw data in FCS Express. All data files, regardless of the scale that they were acquired with, can be displayed in any of the scalings supported by FCS Express.
Biexponential scaling is frequently used to analyze cytometry data because it is capable of displaying data that falls below zero. When using Biexponential scaling, FCS Express automatically calculates the Below Zero Parameter (i.e. the transition point) and the Resolution Minimum to display on a plot from the raw data when the Automatic checkboxes are selected. All data below the transition point value will be scaled as linear, and all data above the transition point value will be scaled as logarithmic.
The ability to automatically adjust plot axes based on the data displayed on the plot aims to avoid potential unexpected negative populations (usually cause by compensation/acquisition issues) that may be missed.
Due to the variability of the raw data (min and max values for the data sets which sometimes can include extreme outliers), the automatic transition point and resolution minimum used to display biexponential scaling on the axis of a plot may change between data files. In FCS Express, it is possible to set both the Below Zero Parameter and define the Resolution Minimum to display on the axes to "lock" the Biexponential scaling so axis do not appear to shift when working between different data files that could have vastly different settings when using "automatic" mode.
How to "lock" the Biexponential Transition Point and Axes on a Plot
In FCS Express, the biexponential scale for a given plot can be locked in place using a combination of the Axis formatting dialog and the Specific Options dialog.
|1. Double click on the plot to open the Format window or right click on the plot and select Format.|
|2. Navigate to Axes using the dropdown menu at the top of the Format window.|
|3. Uncheck the Automatic box beneath Scale.|
|4. Select the Use the Value radio button in the Below Zero Parameter section.|
|5. Adjust the value via the slider bar or by manually entering a value into the text box.|
|6. Uncheck the Automatically set the minimum value check box
*Note: The above steps will need to be repeated for the Y axis.
|The Biexponential transition point where the data goes from linear to log will now be fixed and not change between data files.
Since the default behavior for FCS Express is to automatically determine the Resolution Minimum value within a plot, we will also need to change to this a fixed value.
|7. In the Format window, use the drop down menu to navigate to Specific Options.|
|8. In the Specific Options section, uncheck the box next to Automatic Minimum beneath Resolution Options.|
|9. Insert a value appropriate for the data in the text box next to Resolution Minimum.
*Note: Steps 7 and 8 will need to be repeated for the Y axis.
|The Axis Scaling and Resolution Minimum displayed on the axes will now be "locked" and will not change between data files.|