CytoSwarm – Flow Cytometry Analysis Using uMAP, FlowSOM, TSNE, and More…

 

Find out how to run concatenate, FlowSOM, tSNE, UMAP and more algorithms in CytoSwarm, to interpret your flow cytometry data the way you want to.

 

 

Welcome to the Applied Cytometry algorithm scheduling software CytoSwarm, which performs flow cytometry analysis using UMAP, FlowSOM, tSNE, and many other algorithms to help you interpret your FCS data.

This tutorial has been made to introduce you to CytoSwarm and help you get started using the software.

If you would like to know more please visit the website here, and you will also find links to the installation guide and the user guide here under CytoSwarm.

CytoSwarm is a windows-based application that links with a cloud-based algorithm engine.

Using this system we can run our algorithms on FCS files remotely.

The system is easy to install and easy to use with no knowledge of the r programming language and no local installation of r required.

Using the system, we can sequentially process FCS files through algorithms such as FlowSOM, tSNE, UMAP, and more.

Output files from this system can contain multiple algorithm results when using the system for the first time.

After installation select the Settings button. In the settings dialog you can set up the source folder which will be used for the files you’re going to load.

You can set the download folder location which sets the default folder for downloading files.

This can be modified when saving the files themselves.

If you have not run the software before, you will need to request a license.

To do this, you need to fill in your email address, and a voucher code if you have one, then select the “Generate license request”, which will export a license request file for you to send to info@appliedcytometry.com.

From this, we can generate your license file or key and will send it back via email for you to upload into the software.

In order to add files for processing, press the add files button.

Here you can choose which files you want to process and then press open you can give the job a name, and in this dialog you can also add files or remove files that you wish to.

You can also reorder them using the move up and move down buttons at the side.

Select next when you’re happy with your list and this is where you can compile the list of algorithms that you want to perform on your files, and in the order in which you want them.

Click on the algorithm drop down menu to select the algorithm you want, and press the add button.

For example, if I want to merge all my files I will select merge and then add, and this becomes my step 1.

I can then choose FlowSOM and that becomes my step 2, followed by tSNE which becomes step 3, optimise tSNE, and then UMAP are also added to the end of my list.

cytoswarm Flow Cytometry Analysis Using uMAP, FlowSOM, TSNE, and More...

If you need to change the order in which these algorithms are performed, you can click to move them up or down the list using the buttons at the side.

You can also remove the algorithm if needed later.

We will be able to save this sequence of algorithms as a method, so you don’t need to create a new list and associated settings for each algorithm every single time.

If you have any methods already saved you will be able to use the open method button at the bottom of the dialog to select your preferred method.

In this case we don’t have any saved methods so we are going to step through each of these manually.

If we click next, this dialog shows the parameters in all the files which must be the same.

If they are different the software will give you an error warning and you will not be able to process the files.

Here we can select or deselect parameters you wish to process within the algorithm and these settings will be applied to all files.

Select next and we now step through each of the algorithms in turn.

If you want to use the default parameters for everything, then you can just click the finish button.

Here in this process we will step through each of the algorithms under just the hyper parameters in each.

So, first we have the merge algorithm and then FlowSOM.

Further details on the algorithm are available on the link provided within the software.

If you wish to change the parameters that you are processing, you can click on the channels value and select the parameters you require.

The values within this table can also all be modified.

When we press next, we move on to the third algorithm and so on.

This dialogue at the end shows you the method you have created and the settings within that method, and it is on this page that you can save the method for future use.

Select the finish button and this will upload the files and begin processing for you.

Once all of the files have been sent to the server, then processing can begin.

You can monitor processing by clicking on the job line and then clicking the details button.

In this dialog you will see each of the steps and the progress through each of these steps, so if I click on the merge we can see all of the files that have been merged and that the file save has been completed at the end.

If I click on the FlowSOM we can see no details as yet because it is still preparing.

At any point you can click refresh to update the details held within the preview panel above.

At this point all of the processing is being done on the server and the local application is not required.

You can even close down the software or close down your PC.

Here you can see the completed steps which all have an output FCS file.

You can preview the steps by clicking on them, and a graphic will show off the output if available, and the processing steps will be shown underneath.

Once a job or a series of jobs are completed the status will now show as complete.

You can then download all of the FCS files and the associated images from the server by selecting the item you want to download.

Click the download drop down menu, select in the all steps option will download the original files and all the output files from all the steps together with any previews or images that are created by the algorithm.

If you select results only the system will download just the final output file that has all of the added parameters in.

I will select all steps and select a suitable folder for the download to go into.

After downloading in your target folder, you will find a copy of the original files and each step listed as a file.

Within each file you will have the output FCS file and also any preview images or any other information that’s available within the FCS file.

The original data with the new parameters appended can now be opened for further analysis.

Once you have downloaded all the data that you require, highlight the job and click remove to delete the item from the server.

For more information, visit the CytoSwarm Product Page.

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