Contents
1. So, now you've Clustered... what next?
2. How to Create a Features Special X-Axis Plot
3. Next Steps 
4. Video Overview

So, now you've Clustered... what next? 

Once you have performed Clustering in OMIQ you are left with the task of identifying and understanding the phenotype of these clusters. 

Traditionally this has been done using 2D or 1D plots or Heatmaps and relying on previous manually gated populations. 

Within the Figure Task in OMIQ, you can create a Special X-Axis plot to view several (or all) parameters at once, this overview allows to visualise several markers and assign an annotation or phenotype to your cluster. One advantage of this plot type over a heatmap is you can view biomodal populations rather than making decisions using just an average. 

How to create a Features Special X-Axis Plot

Create a Figure task below your Clustering task and the task where you have added your cluster gates. 

Position in the Workflow for the Figure Task. 

• Create a Scatter plot (overlay)
• Then in Plot Settings, For Overlay type: select Both
• Select Data by Filename and select the files you wish to concatenate together
• For overlay filters, select one broad filter like (all cells or unfiltered) and one of your Clusters

 
 
Plot Settings for the Scatterplot (Overlay)

• Further down in Plot Settings, under Special X-Axis Mode, select Features
• Within Features for X, select the parameters you wish to include
• Note: the scale for the Y axis in non-specific but will be controlled by the Y-axis set at the first Y-axis selection, so we suggest using a fluorescent channel.
  The features will also be added to the plot in the order you select them, so you can control the order of the X-Axis
• Line Width will decide how much your channels blend into one another
• Distribution type will switch between rounded and box channels
  

Plot Settings for the Special X-Axis Plot

In the Plot Settings, you can click to unlink the Resolution, it is then possible to specify and particular X and Y resolution for the optimal size of your plot.  

Unlink your plot Resolution 

You have then made a plot for one filter. 

A Special X-Axis plot looking at one cluster

If you Permute this plot by Filter, you can then select the other cluster filters and have all of your cluster have their own plot. 

Permute your Plot by the filters you are interested in viewing

We suggest adding each plot as a new row. 

Permute using New row

This will produce a plot for each of your clusters, you can review these to help understand the identity of all of your clusters!

Examples of some Overlays looking at Clusters
 

Next Steps 

Once you have understood the phenotype of your clusters, the next step is up to you. You could Rename the cluster with your newly decided phenotype inside the Gating task.

If you want to preserve the original clustering numbers, you could Dupe your Gating task where the Cluster filters were added, and have a new gating task for all your annotated clusters. 

Video Overview