SPADE is performed in FCS Express through the Transformations navigator.

The steps below outline the process of creating a new SPADE transformation for your data:

 

1. Open the Transformation navigator by selecting the Tools tab → Transformations → Transformations command (Figure 9.158, Step 1). The Transformations navigator will open as in the figure below.

2. Click on the drop down arrow adjacent to the blue plus button (Figure below, Step 2).

 

3. Click on SPADE in the drop down list  (Figure below, Step 3).

 

 

The SPADE dialog will now appear in the Transformation navigator as seen in Figure 9.159.

 

 

 

The new SPADE transformation is named as New SPADE by default, but it can be easily renamed by any of the following methods:

•Right click on the SPADE transformation that needs to be renamed→select Rename.

•Select the SPADE transformation that need to be renamed via left click and press F2 on your keyboard.

•Select the SPADE transformation that need to be renamed via left click and click on the button in the Transformations tool bar.

 

1. Choose a Template File for the SPADE transformation by clicking on the ellipsis to the right of the Template File text box (Figure 9.160).  

 

   The SPADE dialog will be populated with a list of available parameters to perform SPADE based on available parameters from the template file (Figure below).

 

 

2.Choose the parameters to be used in the SPADE transformation by checking the boxes next to the parameter names (Figure 9.161, Point 1). Parameters can be filtered or sorted to assist in the selection of parameters when multiple parameters are available in the template file. By right clicking in the parameters section, you can use Sort Ascending, Sort Descending, or Unsorted to easily manage parameter ordering and facilitate parameter selection.  In the right click menu, you can easily select parameters by utilizing Check All, Uncheck All, Check Selected, Uncheck Selected, Invert Selection on All. The options Check Selected and Uncheck Selected allow for using the shift key or Ctrl key to multi-select parameters and check or uncheck them all simultaneously.

 

3.Select or change to the appropriate Parameter Scaling Options for each of the selected parameters if required (a scale of interest can be applied to All or Checked parameters) (Figure below, Point 2).

 

Options can now be set to customize the transformation.

 

 

 

4.(Optional) Select a gate from the Gate drop down menu to only run SPADE on the events within that gate (Figure below, Point 3).

 

5. (Optional) Modify the Display Name Suffix by entering information in the adjacent text box (Figure below, Point 4).

 

6.(Optional) Select a Downsampling Method (Figure below, Point 5). Although multiple Sampling Methods are available (please refer to the Sampling Options chapter for more details), the default Downsampling Method for a SPADE transformation is Target Density (see above for more details and default settings).

 

7. Enter value for Number of Clusters (Figure below, Point 6).

 

8.(Optional) Change the value for Min Percent of Cells Per Cluster (Figure below, Point 7).

 

9.Enter value for Maximum Iterations (Figure below, Point 8).

 

10. (Optional) Change the value for Number of Clustering Attempts (Figure below, Point 9).

 

11. (Optional) Select to Keep Non-Converged Data (Figure below, Point 10).

 

12.Select the Graph Layout Method to be used for the SPADE tree (Figure below, Point 11). Possible choices are Arch (default) or Radio.

 

10. (Optional) Change the Random Seed used for SPADE calculation by either clicking on the Change Random Seed button or by manually inserting a seed in the field (Figure below, Point 12).

 

 

 

The SPADE transformation has now been defined and will be automatically calculated as soon as it will be applied to a heat map. In addition, SPADE will be automatically recalculated as soon as an option in the SPADE dialog will be modified.

 

Given that SPADE is a demanding algorithm that can take several seconds to be calculate, FCS Express allows the user to disable the automatic calculation of SPADE upon options changing and to manually calculate SPADE only when required. To defer calculating the SPADE algorithm, uncheck the box for Automatically Calculate SPADE. Doing so will present you with the button to Calculate SPADE (Figure 9.162) which can be pressed to calculate on demand.

Please note that this option only regulates the auto update of SPADE upon changes made within the SPADE dialog. If SPADE is calculated on a gate and that gate is moved or resized, SPADE will be automatically recalculated regardless this option. If you want to avoid SPADE auto update upon gate moving/resizing, please refer to the Live Updating Gate chapter of this manual.

 

 

 

 

For visualization of the SPADE transformation, a Heat Map may be inserted and modified (Figure 9.163 below). Please also see the Working with SPADE transformations section for additional details on applying SPADE transformations to plots.