Written By: Tim Bushnell, Ph.D.
As a follow-up to our post on tSNE where we compared the speed of calculation in leading software packages, letâ€™s consider the case of SPADE (Spanning-tree Progression Analysis of Density-normalized Events). A favored algorithm in the flow cytometry community, SPADE is used when dealing with highly multidimensional or otherwise complex datasets. Like tSNE, SPADE extracts information across events in your data unsupervised and presents the result in a unique visual format.
Unlike tSNE, which is a dimensionality-reduction algorithm that presents a multidimensional dataset in 2 dimensions (tSNE-1 and tSNE-2), SPADE is a clustering and graph-layout algorithm. The ...
Written By: Tim Busnell, Ph.D.
1. How it works
2. Panel design
3. Sample preparation
4. Data analysis
5. Imaging mass cytometry
Todayâ€™s article will summarize the functionality of mass cytometry technology. This tech has been commercialized largely by Fluidigm in the CyTof systems. There are 5 key points to cover, or takeaways, that cytometrists should keep in mind as they perform their research.
How Does Mass Cytometry Work?
Traditional fluorescent flow cytometry has started to push the limit of the number of simultaneous parameters that can be measured. With the recent advent of spectral cytometry, as many as 40 simultaneous fluorescence parameters can be measured.
The first foray ...
Written By: Heather Brown-Harding, Ph.D.
When you need better resolution than what can be achieved using a traditional microscope, it can be very intimidating to figure out which microscopes will work best for your experiment. Super-resolution imaging methods require software reconstruction after image acquisition. This is because multiple images are acquired, and they need to be combined. Additionally, the points of light need to be reassigned to their true location.
Today, we're going to discuss 5 different super resolution methods their pros and cons. Although Rayleigh Criterion is not broken, these techniques each feature creative ways to get around it.
1. Structured illumination ...
Written By: Tim Busnell, Ph.D.
Reproducibility is key to the scientific method. After the results of a study are published, the community validates the findings and extends them. If the findings are not reproducible, the second step is impossible. With performable experiments increasing in complexity, and the concurrent increase in the cost of equipment and reagents to perform these experiments, it is important to find the best way to maximize the money spent on advancing research.
In flow cytometry, there are many places where improvements can be made to increase the consistency and reproducibility of an experiment. The most obvious place is ...
Written By: Tim Bushnell, Ph.D.
All flow cytometer instruments have a certain 3 components you likely donâ€™t know enough about. The way they are put together will dictate the performance of the system. As a user, youâ€™ll be interacting heavily with these components, so you need to know both what they are and how they work.
The 3 components are the fluidics, optics, and electronics. Fluidics will be managed in order get interactions at the proper flow rate, ensuring that your data keep a tight CV. After cells have passed the laser intercept, you need a way of measuring the signal. This ...
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MedImmune purchased a site license of Expert Cytometryâ€™s Mastery Class. We have over 100 scientists using the program and the feedback has been phenomenal. Iâ€™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. Iâ€™m now able to focus my attention, and that of my staffâ€™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.Â 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, ...
Expert CytometryÂ 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. Â Â
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.
I was (and still am for the most part) completely ignorant of the details of flow cytometry.Â The Expert Cytometry teamÂ 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 CytometryÂ responded and completed the analyses when we were on short timelines. We continue to work with ExCyte and will for the foreseeable future.
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 canâ€™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 weâ€™ll bring ExCyte back for another course in ...