Open Flow Cytometry Videos

Open Flow is an open access Flow Cytometry education resource. Live interactive classes are held monthly and are free for all who wish to register. The Open Flow initiative is a collaboration between the Francis Crick Institute (London, England), Memorial Sloan Kettering Cancer Center (NY, NY, USA) and the Whitehead Institute for Biomedical Research (Cambridge, MA, USA).

Questions? Email openflowcytometry@gmail.com

Flow Cytometry Home | Resources | Rates | Instruments | Training & Access | Sample Prep Guidelines & SOPs | Software | Biosafety | Contact | Scheduling for Flow Cytometers 

Videos
Experimental Setup in Diva

Join Open Flow as we explore experimental setup in Diva. During this classroom, we explored the importance of flow cytometer instrument configuration, laser delays, QC via CS&T, basic overview of BD FACSDiva software and setup of a basic one color flow cytometry experiment with lyopholized cells stained with a single fluorophore.

Experimental Voltage Optimization in Diva

When setting up a flow cytometry experiment, voltage optimization is required to ensure you are resolving your populations of interest to the best of your ability. In this Open Flow session, we will review in depth how to carry out a voltration of single color cells and how to determine optimal voltages through statistical analysis. Diva acquisition software tools and tricks will also be reviewed.

Fluorescence Compensation in Diva

When carrying out multicolor Flow Cytometry experiments, it is common to experience fluorescence spillover due to overlapping emission. In this Open Flow session, we will review in depth the principles behind compensation and how it is carried out on a BD Fortessa analyzer. This will be done utilizing the automatic compensation built into the software and a four color experiment.

Multicolor Troubleshooting in Diva

 

When carrying out multicolor flow cytometry experiments, you may run into scenarios where your data does not look as expected. In this Open Flow session, we will review in depth the rules of compensation and how our data can suffer when we do not follow them. This will be done utilizing the automatic compensation built into the software and a four color experiment.

Full Spectrum Flow Cytometry with the Cytek Aurora

 

In this OpenFlow session we will review the theory and practice of full spectrum flow cytometry. We will address some of the fundamentals behind full spectrum flow cytometry and how it compares with the current technology, as well as running a multiparameter experiment on a five-laser Cytek Aurora using Spectroflo software.

 

Full Spectrum Flow Cytometry Part II - Panel Design & Troubleshooting

In this session, we will continue to look at full spectrum flow cytometry. Specifically, we will review considerations for fluorescence panel design. What do we have to think about to design a successful panel and how can we combine fluorochromes that might be difficult on a conventional flow cytometer? We will also look at autofluorescence extraction and present some troubleshooting tips. The session will be a mixture of theory and practice using a 5 laser Cytek Aurora.

Introduction to Cell Sorting Part I

In this session, we will explore cell sorting by flow cytometry. This is an extremely useful technique as it allows us to separate cells of interest for further study. However, the success of a sort depends on many factors including the sample preparation and knowledge of how the sorter works. In part I, we will look at the theory of cells sorting and perform a simple sorting experiment using a Sony SH800 cell sorter.

Introduction to Cell Sorting Part II

In this OpenFlow session we will review the theory and practice of cell sorting. We will consolidate knowledge from our previous session and use a BD FACSAria to perform a more complex cell sort. The aim of this session is to show attendees the theory of cell sorting in addition to the practicalities of performing a successful sort.

Introduction to Cell Sorting Part III

The success of a cell sort depends on many factors including the sample preparation and knowledge of how the sorter works. In this session, we will look at what can go wrong and see the importance of experimental optimization. Delegates will also discover how to troubleshoot the most common problems and will have the opportunity to ask questions.

Note: At the end of this session an issue was seen with post sort purity of populations for the four-way sort. After the session, we re-checked drop charge delay, which was unchanged, and a repeat sort was performed on two separate Arias which yielded expected results

Introduction to Flow Cytometric Data Analysis Part I

In this session, we looked at data analysis of flow cytometry files. In this first session presented an overview of data analysis and how it can be used to answer the biological questions you have using FlowJo software. This included quality control, choice of plots, regions and gating, data scaling and selection of appropriate metrics.

Introduction to Flow Cytometric Data Analysis Part II

In this second session we will look in more detail at how we can report metrics to answer biological questions which will include looking at statistics, the proper use of controls and how to display your data. In this session we will use FCSExpress software. We will run this analysis live and interactively allowing delegates to ask questions as we go along.

Introduction to Flow Cytometric Data Analysis Part III (Compensation)

In this third session we will look in more detail at offline compensation using both FlowJo and FCSExpress to show manual and automatic compensation and how to assess success. We will run this analysis live and interactively allowing delegates to ask questions as we go along.