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Ensuring Reproducibility In Flow Cytometry

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Planning For Surface Staining Of Cells In Flow Cytometry

Written by Tim Bushnell, Ph.D One of the most common assays in flow cytometry is the surface labeling of cells with antibodies. Often termed “immunophenotyping”, it allows the researcher to identify, count, and isolate cells of interest in a mix of input cells. Every lab has their own favorite protocol, handed down from some hallowed, chemical-stained notebook, and followed as exactly as making a souffle. The real questions are, which of those steps are critical, and (with changes in instruments and theory) what other factors should be considered when staining cells? This article will focus on staining immune cells, but the principles ...

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2 Key SPADE Parameters To Adjust For Best Flow Cytometry Results

Written by Tim Bushnell, Ph.D Mass cytometry panels routinely include 30 or more markers, but traditional analysis methods like bivariate gating can’t adequately parse the resulting high-dimensional data. Spanning-tree progression analysis of density-normalized events (SPADE) is one of the most commonly used computational tools for visualizing and interpreting data sets from mass cytometry and multidimensional fluorescence flow cytometry experiments. There are two key parameters in SPADE that you can adjust in order get the best results possible: downsampling and target number of nodes, or k. Knowing how to properly set these values will enable you to enhance the quality of your analysis. Downsampling Imagine your ...

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3 Advantages Of Using The ZE5 Cell Analyzer

Written by Tim Bushnell, Ph.D Since the first laser was mounted to create the first flow cytometer, there has been a push for more - more lasers, more detectors, and more colors. Why? So researchers could ask more complicated questions to squeeze every iota of data out of rare events and precious samples, and so clinicians could expand the diagnostics capabilities of the technology. In addition, this trend has occurred so biotech companies could expand high-content screening for drug discovery. Instrument manufacturers have brought to the table a plethora of different instruments, with capabilities to suit the needs of the researcher, at price points ...

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The Difference Between Linear And Log Displays In Flow Cytometry

Written by Tim Bushnell, Ph.D Data display is fundamental to flow cytometry and strongly influences the way that we interpret the underlying information. One of the most important aspects of graphing flow cytometry data is the scale type. Flow cytometry data scales come in two flavors, linear and logarithmic (log), which dictate how data is organized on plots. Understanding these two scales is critical for data interpretation. Let’s start at the beginning, where signal is generated, and trace its path all the way from the detector to the display. Behind every flow cytometry data point is what we call a pulse. The pulse is ...

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4 Ways To Achieve Reproducible Flow Cytometry Results

Written by Tim Bushnell, Ph.D Reproducibility is one of the hallmarks of the Scientific Method. One reason scientists publish their results is so that other labs can attempt to reproduce the results and extend the investigation into new areas. From the Begley and Ellis commentary in Nature, to the development of the Rigor and Reproducibility initiative at NIH, to articles in the mainstream media, reproducibility is on everyone's mind. This is not a bad thing, and following the best practices in flow cytometry provides investigators, peer reviewers, and colleagues with more confidence in your data. There are several areas within the process of ...

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Testimonials
MedImmune purchased a site license of Expert Cytometry’s Mastery Class. We have over 100 scientists using the program and the feedback has been phenomenal. I’m amazed at how much the scientists are coming to me with questions that are much more advanced than what I used to get before having access to the Mastery Class. I’m now able to focus my attention, and that of my staff’s, to more advanced questions and inquiries specific to the researcher. We are also having less trouble with the mistakes that new users make and spending less time dealing with troubleshooting basic issues. As the core director at MedImmune, I definitely recommend the Expert Cytometry Mastery Class to all cores, particularly in a corporate setting, where we simply do not have the time to train all of the new users in best practices. My core is running 10 hours a week, ...

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Johnna Wesley, Ph.D.Senior Manager, Novo Nordisk
Expert Cytometry provided an excellent course to our group. The course was over two days. On day one, the instructors provided a great overview of core concepts of flow. On day two, we were able to customize the course to our needs. It was very valuable and I would recommend that if your group is starting to use flow more heavily, that you bring Expert Cytometry in for a course. There are nuances that I never knew and it immediately helped our group perform better experiments.   

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Before taking this course, my experience with flow was limited to simple 2 color experiments. Then I was asked to perform a 10-color complete immunophenotype experiment in non-lymphoid tissues. With a ton of help from a long time co-worker and friend, I was successful with the effort. She highly recommended the Excyte Mastery Course to me and I can say she was right. This course truly elevated my knowledge and understanding of flow cytometry. It was very organized, concise and covered everything from understanding your instrument, optimizing it's performance, designing multicolor panels and executing a flow cytometry experiment with all the necessary controls. I look forward to learning more from the advanced seminars and webinars. I highly recommend this course.

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I was (and still am for the most part) completely ignorant of the details of flow cytometry. The Expert Cytometry team provided explanations and helped me understand the technology to the point where I was able to intelligently and efficiently manage both of our contract flow labs. Expert Cytometry provided independent assessments of the quality of the work and data from the labs that allowed me to make good decisions regarding the results. Finally, I was always amazed at how quickly Expert Cytometry responded and completed the analyses when we were on short timelines. We continue to work with ExCyte and will for the foreseeable future.

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Our group at BlueBirdBio was relatively new to flow. We brought ExCyte in for a basic course, and also to help train us on best practices for our new flow cytometer. The level of flow cytometry knowledge ExCyte brought with them was amazing. I can’t believe how much experience ExCyte has and how much they bring to the courses they provide. ExCyte was able to answer all questions with real world examples. The course brought the theory of flow to the real world and helped us understand the concepts more easily. ExCyte was professional, courteous, and it almost felt like they became part of our group while we were working together. I know I can ask ExCyte any questions whenever I need them and I hope to continue to work with them as we move forward. I have no doubt we’ll bring ExCyte back for another course in ...

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