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Expert Technical Training That Ensures Reproducibility In STEM

Cheeky Scientist’s Technical Programs currently include Expert Cytometry, Expert Microscopy, and Expert Sequencing. Each program provides continuing education credits and is taught by leading PhD experts in their respective STEM fields.

Expert Technical Training That Ensures Reproducibility In STEM

Top Technical Training eBooks

Get the Advanced Microscopy eBook

Get the Advanced Microscopy eBook

Heather Brown-Harding, PhD

Learn the best practices and advanced techniques across the diverse fields of microscopy, including instrumentation, experimental setup, image analysis, figure preparation, and more.

Get The Free Modern Flow Cytometry eBook

Get The Free Modern Flow Cytometry eBook

Tim Bushnell, PhD

Learn the best practices of flow cytometry experimentation, data analysis, figure preparation, antibody panel design, instrumentation and more.

Get The Free 4-10 Compensation eBook

Get The Free 4-10 Compensation eBook

Tim Bushnell, PhD

Advanced 4-10 Color Compensation, Learn strategies for designing advanced antibody compensation panels and how to use your compensation matrix to analyze your experimental data.

Technical Training Programs

Technical Programs
Expert Cytometry

Expert Cytometry

Get access to expert training in flow cytometry, including real-time guidance by top flow cytometry scientists.

Expert Sequencing

Expert Sequencing

Get access to expert training in next generation sequencing (NGS) by top NGS experts.

Expert Microscopy

Expert Microscopy

Get access to expert training in microscopy, including daily mentorship by microscopy experts across disciplines.

Featured Articles

How to Perform Doublet Discrimination In Flow Cytometry

How to Perform Doublet Discrimination In Flow Cytometry

By: Tim Bushnell, PhD

You are probably familiar with the term, “doublet discrimination” or “doublet exclusion”, and have likely included this flow cytometry measurement into at least some (if not all) of your gating strategies. Even though you may utilize this important gating strategy, you may not have had the chance to delve deeper to explore exactly what doublets are and why it’s critical to exclude them. This article aims to give you insight on the what, why, and how of doublet discrimination.

What Is A Fluorescence Minus One, or FMO Control

What Is A Fluorescence Minus One, or FMO Control

By: Tim Bushnell, PhD

The Fluorescence Minus One Control, or FMO control is a type of control used to properly interpret flow cytometry data.  It is used to identify and gate cells in the context of data spread due to the multiple fluorochromes in a given panel. An FMO control contains all the flurochromes in a panel, except for the one that is being measured.  For example, in the four color panel, there would be four separate FMO controls, as shown in the table below. The FMO control ensures that the any spread of the fluorochromes into the channel of interest is properly identified.…

How To Analyze FACS Data And Prepare Flow Cytometry Figures For Scientific Papers

How To Analyze FACS Data And Prepare Flow Cytometry Figures For Scientific Papers

By: Tim Bushnell, PhD

When preparing figures for publication, the scientific question and hypothesis that forms the basis of the paper must be central and all the figures must be in support of that. The flow cytometry data that forms the basis of the conclusions should be presented clearly and concisely. While it provides pretty pictures and colorful layouts, the meat of the data are the numbers ― percentages of populations, fluorescent intensity levels and the like ― are what will convince the reader that the hypothesis tested is valid and well thought out. Here’s how to choose the correct flow figure for presenting your data.

5 Essential Calculations For Accurate Flow Cytometry Results

5 Essential Calculations For Accurate Flow Cytometry Results

By: Tim Bushnell, PhD

Flow cytometry is a numbers game. There are percentages of a population, fluorescence intensity measurements, sample averages, data normalization, and more. Many of these common calculations are useful, but surrounded by misconceptions. This primer will help you decide which calculation to use, when to use it, and how to interpret the results.

When To Use (And Not Use) Flow Cytometry Isotype Controls

When To Use (And Not Use) Flow Cytometry Isotype Controls

By: Tim Bushnell, PhD

The field of flow cytometry is moving beyond the use of isotype controls, with many suggesting they be left out of nearly all experiments. Yet, isotype controls were once considered the only negative controls you should ever use. They are still very often included by some labs, almost abandoned by others, and a subject of confusion for many beginners. What are they, why and when do I need them? Are they of any use at all, or just a waste of money? Most importantly, why do reviewers keep asking for them when they review papers containing flow data? Here is everything you need to know about using (or not using) isotype controls in your next flow cytometry experiment.

3 Flow Cytometry Gates That Will Improve The Accuracy Of Your FACS Data Analysis

3 Flow Cytometry Gates That Will Improve The Accuracy Of Your FACS Data Analysis

By: Tim Bushnell, PhD

When training new users on data analysis, there are several different best practices and gating strategies you should incorporate into your analysis. There are also several misconceptions you must understand. There are 3 gates that many researchers are not using but should be using when analyzing their flow cytometry data. These gates are critical for good data analysis. They will help remove many confounding events that may be clouding your analysis, especially where rare events are concerned.

How To Set And Monitor Optimal Voltages For A Flow Cytometry Experiment

How To Set And Monitor Optimal Voltages For A Flow Cytometry Experiment

By: Tim Bushnell, PhD

The best way to take out the fear and agony of setting voltages is to use some optimization methods. The peak 2 method is a useful and robust method of identifying optimal PMT voltage ranges. Refining that to the voltage walk with the actual cells and fluorochromes of interest will further improve sensitivity, which is especially critical for rare cell populations or emergent antigens. This article describes how to set up, monitor, and maintain optimal voltage settings for your flow cytometry experiment.

5 Gating Strategies To Get Your Flow Cytometry Data Published In Peer-Reviewed Scientific Journals

5 Gating Strategies To Get Your Flow Cytometry Data Published In Peer-Reviewed Scientific Journals

By: Tim Bushnell, PhD

When sitting down to perform a new analysis of flow cytometry data, the researcher is guided by very particular laws of nature and a very specific method of working through a biological hypothesis to avoid shaping the results to his or her whims. Following these 5 data analysis and gating strategies through the hierarchy described in this article, researchers are provided with several strategies for identifying and displaying the most relevant data from their flow cytometry experiments.

Why You Need To Use FMO Controls For All Multicolor Flow Cytometry Experiments

Why You Need To Use FMO Controls For All Multicolor Flow Cytometry Experiments

By: Tim Bushnell, PhD

FMO controls are samples that contain all the antibodies you are testing in your experimental samples, minus one of them. When analyzing the minus, or left out parameter in an FMO control, you give yourself a strong negative control to work with. It’s a strong negative control because the left out marker in the FMO control allows you to take into account how the other stains in your panel affect the respective minus parameter. Many flow cytometry gates are difficult to define. This is especially true when you’re looking at activation markers within a continuum or accounting for the large data spread that occurs when compensating a 10+ color experiment. The only way to convince reviewers that your gate is in the proper place is by using FMO controls. Here's why you need to use FMO controls for any multicolor flow cytometry experiment and how to prepare these controls properly.

What Is Flow Cytometry Light Scatter And How Cell Size And Particle Size Affects It

What Is Flow Cytometry Light Scatter And How Cell Size And Particle Size Affects It

By: Tim Bushnell, PhD

Forward scatter detectors collect light at small angles relative to the incident beam and can take advantage of the fact that cells preferentially scatter light in this “forward” direction. Forward scattered light is traditionally and often effectively measured with a photodiode, rather than the more sensitive photomultiplier used to measure fluorescence and side scatter. Scatter gets dim very quickly when particles have diameters below the wavelength of illuminating light, considering that scatter intensity decreases with a dependence on r6 of the particle. Here’s how small particles affect light scatter.

4 Biggest Mistakes Scientists Make During Multicolor Flow Cytometry Cell Sorting Experiments

4 Biggest Mistakes Scientists Make During Multicolor Flow Cytometry Cell Sorting Experiments

By: Tim Bushnell, PhD

Multicolor sorting experiments can be complicated and if not setup properly, result in wasted time and suboptimal results. When setting up a multicolor experiment, the most saliently critical step is to set PMT voltages properly. In addition, using a viability dye and addressing doublet discrimination and setting the right sort regions and gates is important for any kind of flow cytometry experiment, but particularly for cell sorting. These tips help to ensure your setup is perfect to achieve results of the highest caliber.

What Is Sheath Fluid

What Is Sheath Fluid

By: Tim Bushnell, PhD

Sheath fluid is the solution that runs in a flow cytometer.  Once the sheath fluid is running at laminar flow, the cells are injected into the center of the stream, at a slightly higher pressure.  The principles of hydrodynamic focusing cause the cells to align, single file in the direction of flow. Depending on experimental needs, different formulations of sheath fluid can be used. Many labs purchase pre-mixed phosphate-buffered saline from Leinco Technologies. Some researchers use Hepes-buffered saline.  This is particularly useful for high-pressure cell sorting as Hepes controls pH better at high pressure than phosphate buffers do. Finally, since…

Success Stories

Fabienne Lucas, MD, PhD

Fabienne Lucas, MD, PhD

Brigham and Women's Hospital
Resident in Clinical Pathology

I began Expert Cytometry as a postdoc in the US after my medical and PhD training in Germany and the UK. I had used flow cytometry for more than 5 years but needed to take a ‘deep dive’ to really understand the technique. I soon became the expert in my lab and helped my peers design their experiments. It was also the impetus for me to apply (and receive) the Mary Lou Ingram Scholarship, and start the MiSET RFC standard while I continue my career in cytometry.

Carina Torres

Carina Torres

Core Manager
Eli Lilly & Company

I invited ExCyte to Lilly Pharmaceuticals to teach a two-day comprehensive course; ranging from the fundamentals of flow to the experimental design of assays specific to what I think a lot of biotech companies are doing these days. Afterward, several of our scientists wondered why I didn’t bring in a course like this earlier! Our instructor, Tim Bushnell, Ph.D., educated my users with the background they needed to design their assays most effectively. I immediately noticed the positive impact he had when my users began to ask more informed questions during the planning stages of their experiments. As a busy core manager, it’s difficult to find the time to prepare such a well-designed course like ExCyte offers while trying to stay focused on innovation and maintaining a competitive advantage in the fast-paced industry environment. I’m grateful to the ExCyte team for helping me to keep things moving forward at Lilly.

Amrutesh Puranik, PhD

Amrutesh Puranik, PhD

Postdoctoral Fellow
Mayo Clinic

I’m a postdoctoral researcher and know a lot about flow but I needed to learn a few new techniques and execute them before I could get my paper published. I joined Expert Cytometry because they have a good reputation for teaching high-level flow cytometry information very well. I really enjoyed the personal guidance the Cheeky Scientist Technical Program offered and was impressed by how much I learned about controls and compensation, experimental design, statistics, and even apoptosis. Cheeky Scientist's ExCyte is perhaps the only class that teaches you from scratch how to be a flow cytometry expert.

Miguel Garcia

Miguel Garcia

Core Manager
Swiss Federal Institute of Technology

As a Core Manager, my position requires staying up to date with the latest best practices. The Cheeky Scientist Technical Programs provided the necessary information on flow cytometry to stay on the cutting edge of the field for a reasonable investment. Overall, the course was really well done by speakers expert on the field and I really appreciated the flexibility of listening to the webinars as needed. The Cheeky Scientist Technical Program is wonderful and I would recommend it to beginner as well as advanced flow cytometry professionals.