Instrument Specific Settings

Restore Default Settings

Restores the list of Instruments and Instrument Specific Settings back to the default set and settings provided with FCS Express.

NOTE: when updating to a new release of FCS Express from an existing release, new ISS that have been added to an update may require the user to restore the default ISS settings to enable the new ISS that were provided in an update. Any custom ISS that you have previously created will need to be recreated after restoring the default settings.

Duplicate

When an instrument specific settings is selected from the list, pressing Duplicate will create a copy of the setting. The copy maybe be renamed in the Instrument Name section and new identification rules and settings may be defined.

Add

The add button will allow you to create your own customized instrument specific setting from scratch. When the Add button is clicked a dropdown menu will appear allowing the choice of:

•General

•Cytek

•Sony

•Merged FCS Files

Choose from one of the options in the list to best match the data source type you intend to create an instrument specific settings for.

Remove

When an instrument specific settings is selected from the list, pressing Remove, will remove the setting from the list. Please note that Default FCS 2.0 and Default FCS 3.0 settings may not be removed.  

Up and Down Arrows

When an instrument specific settings is selected from the list, pressing the Up or Down arrows will move the setting up or down the list. Settings higher in the list will be given priority over those lower in the list.

Instrument Name

Controls the name displayed in the Instrument Specific Settings list for the setting currently selected. Note this may not be changed for Default FCS 2.0 and Default FCS 3.0 settings.

Identification Rule

The instrument origin of data files will generally be recognized by the cytometer ($CYT) keyword, or a formula which evaluates multiple keywords, that are recorded as metadata by the cytometer at the time of acquisition.  Several Instrument-Specific Settings and Identification Rules have been included by default. Identification rules may be edited or added following the syntax with quotes (with $CYT as an example):  "<$CYT>" = "value of the $CYT keyword for your data files".  Operators such as OR, AND, NOT, and other common operators, may also be used. For instance '("<$CYT>" = "Accuri C6") or ("<$CYT>" = "BD Accuri C6")'

Value to start for log parameters

Choose the starting value for parameters displayed in the log scale from the drop-down list.

Keyword To Use as Parameter Name

Choose a keyword (or combination of keywords) to use as parameter display names from the drop-down list.  This option will define how the X and Y axis labels are displayed within a plot.

The options are Name, Stain, Name and Stain, or Stain and Name.  These options are specifically referring to the keyword values for $PnN and $PnS.  These keywords designate the Parameter Name and Parameter Stain keywords written to file by the acquisition software.

For example, if choosing to display the 3rd parameter written within a data file and the value of the Stain and Name keywords are as follows:

•$P3N = FITC - A

•$P3S = CD3

The Name option will display FITC - A.

The Stain option will display CD3.

The Stain and Name option will display CD3 FITC - A.

The Name and Stain option will display FITC - A CD3.

Keyword for Parameter Matching

This option is used when the In new layouts, remember parameters by option has been set to Name and will define the text that FCS Express uses when matching the parameters to be displayed within a plot.

The options are Same as Display Name, Name, Stain, Name and Stain, or Stain and Name.

Please note that the text has to be an exact match between files for FCS Express to properly know which parameter to display when a new data file is loaded into a plot.

For example, if this option is set to Name and your data has a $PnN value of FITC - A, the next data file loaded into a plot needs to have a corresponding parameter with the exact $PnN text match of FITC - A to properly display axis labels.

If this option is set to Stain and Name and your data has a $PnS value of CD3 and a $PnN value of FITC - A, the next data file loaded into a plot needs to have a corresponding match with the exact $PnS text match of CD3 and $PnN text match of FITC - A to properly display axis labels.

When FCS Express attempts to match parameters, for instance, when defining compensation controls, these settings will be also be used to attempt matching.

Merge File Import Options

These options allow to automatically create the File Identifier (i.e. numeric) and/or the Classification (i.e. file names) Columns when FCS files are loaded as Merged FCS File (see the Adding Merged Files to the Data List and the Working with Heat Map chapter of the manual for more details on the Merge FCS File format).

Additional Classification keywords can also be added using any of the keywords available in the loaded files. Multiple keywords can be specified as comma-separated. If a keyword is specified and multiple files share the same value for that keyword, events from those files will be grouped together under the corresponding classification parameter.

Range Options

Default Range Options may be specified for Scatter and/or Fluorescence Parameters. You may choose from four mutually exclusive options:

•Automatic:  FCS Express will set the Min/Max plot Resolution automatically. The Resolution Minimum will be automatically defined by FCS Express based on the selected data Scale (see Scale Options below). The logic for the Resolution Maximum definition varies between data types, but for most flow cytometry data it is set according to the what is specified by the metadata (keywords) for each parameter in the data file.

The Automatic setting is the default setting used by FCS Express if the data file does not meet any of the Identification rules in the list.

•Based on data:  For each plot, and for each parameters displayed on the based overlay of that plot, FCS Express detects the minimum and the maximum of the data, and set the plot Resolution accordingly (a margin will be added so that data does not appear cut off on the plot boundaries). Please note that Scatter and Fluorescence Parameters have independent options and thus can be customized independently.

To enable the Based on data setting, add a checkmark to the box called Minimum and/or Maximum, depending on the one(s) which should be based on data.

•Percentile:  FCS Express will set the plot Resolution according to the specified Percentile.  This is a convenient way to automatically not plot outliers which may located far beyond the data of interest.  In effect, your plots will already be zoomed into your highest-level region of interest.

•Manual: this option allows you to manually specify a Minimum and Maximum for the plot Resolution. The entered values will be used for all parameters and all plots (please note that Scatter and Fluorescence Parameters have independent options and thus can be customized independently).

To enable the Manual option, enter the desired minimum and maximum values in the two fields below "Manual". Enter the minimum in the upper field and enter maximum in the lower field.

Scale Options

When the Automatic check box is checked FCS Express will automatically apply a predefined scale specific for the selected instrument. To manually specify a scaling option, first uncheck the Automatic box then select the default scaling you wish to apply from the drop down menu which will appear.

Division Factor

The Division Factor may be specified for Scatter and/or Fluorescence Parameters by adjusting the numeric value in the Divide Axis Labels Factor text box.  In addition, the user may specify Do not perform scaling, Scale by factor, Scale to a maximum value. These may be specified for both Scatter and Fluorescence parameters.  The Do not perform scaling does not make any adjustments to scaling. The Scale by factor allows for the data to be displayed with a different resolution (# of channels) than its native resolution.  The Scale to a maximum value drop down menu contains common display scales.

Import spectra as individual Parameters

When checked, Spectrum plots will allow the user to select any individual laser spectra and a merged spectra as a parameter, for instance, Violet, Blue, Red and Violet\Blue\Red will all be available. If unchecked, only one merged parameter for all laser spectra will be displayed, for instance, only one parameter for Violet\Blue\Red will be available.

Import spectra as Single Merged Parameter

When checked, a single merged parameter for all laser spectra will be available for selection. For instance, if Violet, Blue, and Red were collected, a new merged parameter for Violet\Blue\Red will be available. If unchecked, only individual spectra will be accessible as long as Import Spectra as individual Parameters is checked (above).

Spectra Parameter Label

The options allows users to define which Spectra Parameter to use as a label on spectrum plots, either Wavelengths or Parameter Names. If Wavelengths are selected, Spectrum plots will support displaying the numeric wavelength on the X axis, if Parameter Names is selected, Spectrum plots will support displaying the parameter names on the X axis, for instance, V1-A, V2-A, V3-A.

Delete fluorescence parameters after spectra merge

If checked, additional spectra parameters (for instance V1-A, V2-A, V3-A) will be removed when visualizing data in 2D and 1D plots. If unchecked, individual spectra parameters will be accessible in 2D and 1D plots at the end of the standard parameter list.

Merge Raw and Unmixed Cytek Files during import

When checked, if raw and unmixed data files exist in the appropriate file structure, and the unmixed data file is loaded into any plot(s), a resulting spectrum plot inserted for the data set will visualize all spectral parameters. If unchecked, when loading an unmixed data file, a spectrum plot for the data set will not be accessible since raw spectral data will not be merged with the unmixed results. When loading a raw data file, only scatter parameters will be accessible on 2D and 1D plots but the raw spectral data will be accessible in a spectrum plot.

Cytek File Merge Rule

Four options are available and described below:

Load User Selection: Uses the check box option set above for "Merge Raw and Unmixed Cytek Files during import"

Load both Raw and Unmixed Always: Loads both Raw and Unmixed into a merged data file for use in 2D, 1D, and spectrum plots regardless of the check box option set above for "Merge Raw and Unmixed Cytek Files during import"

Load Unmixed Only: Loads only the Unmixed version (no spectral parameters) of the data set when loading data.

Load Raw Only: Loads only the Raw version of the data set (only spectral parameters) when loading data.

Spectrum Parameters to Read

The options allows users to define which Spectrum Parameters to Read, and to display on a Spectrum plot, either Wavelengths or Channels or both (one choice must be selected). If Wavelengths are selected, Spectrum plots will support displaying the wavelength on the X axis, if Channels is selected, Spectrum plots will support displaying the Channels on the X axis. If both are selected, Spectrum plots will support displaying both Wavelength and Channels on the X axis.