How Droplets Are Charged And Drop Delays Are Determined During An Electrostatic Cell Sorting Experiment
Written by Michael Kissner.
They way that droplets are charged is one of the more counterintuitive and complex aspects of cell sorting.
My own experience as a past fledgling cytometrist certainly echoes this assertion. I distinctly remember my struggle to understand how exactly all of the sorting components coalesced to accomplish the instrument’s tasks, but when I finally did, I gained a much deeper understanding of cell sorting as a holistic process rather than a vague sum of its parts.
It’s my hope here to be able to facilitate a more accurate conception of the process, grounded in a reality of physics and mechanics rather than one shrouded in magic.
Towards this end, the most crucial point is that the technique of electrostatic cell sorting, which is the most widely employed type of sorting, is fundamentally built upon droplet charging.
How Flow Cytometry Electrostatic Cell Sorting Works
Electrostatic cell sorting is a complicated process that continues to be improved.Read More
How To Use Flow Cytometry To Correctly Define T Cell Subsets And Their Functions
Written by Jennifer Snyder-Cappione, Ph.D.
Flow Cytometry is a remarkably powerful tool for the study of T cells. It has been successfully used for many decades to accurately visualize and enumerate a variety of T cell subsets.
With a large sensitivity range for fluorescent probes, >95% sampling efficiency, and the ability to sort populations of interest for further study, fluorescent-based cytometry remains a tool of choice for T cell analysis.
Single cell visualization of T cells in a heterogeneous sample is clearest when the defined T cell populations are determined with ‘rock-solid’ gating and data analysis strategies.
For example, detection of the total CD4 and CD8 T cell compartments (via CD3+ CD4+ and CD3+ CD8+ cells, respectively) is straightforward; also, T cell populations that are clearly defined by surface antigen expression include antigen-specific (tetramer-binding) memory T cell clones and invariant Natural Killer T (iNKT) cells, a unique T cell subset discerned via binding to a CD1d-glycolipid loaded tetramer.
Such gating strategies, when paired with CD3 inclus ...Read More
What Is The International Cytometry Certification Exam (ICCE) And How To Pass It
Written by James McCracken, Ph.D.
Clinical workers in flow cytometry and other fields have had certification requirements for quite some time.
However, scientists in the basic research space have relied on experience and pedigree as a metric for skill and knowledge, not certifications.
Professional certification is a process where an individual can take some form of exam that demonstrates a level of competence that a body of experts have determined are necessary for performing the job that the certification covers.
This process is also a way to improve best practices in a particular field, such as the field of flow cytometry.
There is now an option for certification in flow cytometry.
This option offers a way for you to help assure potential recruiters that you have a particular knowledge base in the theory and practice of flow cytometry.
This certification process is known as the International Cytometry Certification Examination (or ICCE), which leads to the CCy designation for those who pass the test.
What Is The International Cytometry Certification Exam?
The ICCE was develope ...Read More
How To Detect Microvesicles, Microparticles, And Ectosomes By Flow Cytometry
Written by Phil Hexley, Ph.D.
There seems to be a big fuss about small flow these days.
What is small flow?
Microvescles, microparticles, ectosomes—the terminology is all over the place but one thing is certain…
There is a need for flow cytometric analysis of extracellular vesicles.
To avoid ambiguity in this post, the population of extracellular vesicles discussed here shall be called microvesicles.
These small, membrane bound fractions of cells released during activation and apoptosis are in the 0.1 to 1 micron diameter size range (or there about depending on which article you read).
Released in vast quantities and carrying a variety of traits from the originating cell these microvesicles are known to be involved in a wide range of processed from stem cell renewal and tumor metastases through to coagulation and inflammation, just to name a few!
They key is that microvesicles have a huge potential to be important biomarkers in early disease detection and/or treatment progression.
What’s Down There? Debris Or Microvesicles?
Microvesicles have been known about for a long time, b ...Read More
5 Important Peer Review Questions To Answer Before Submitting Your Flow Cytometry Data
Publish or perish remains the mantra of science.
All the experiments and experience in the world do not count if you are unable to communicate your results to the scientific community.
As part of that communication process, your paper will undergo the dreaded ‘Peer Review’ process. If you wish your paper to survive the peer review process, you must collect, analyze, and present your data properly—before you submit your paper.
A review of the following peer review questions, as well as how to answer these questions, will help ensure your paper is not rejected.
5 Questions Peer Reviewers Will Ask
As one who is asked to peer review papers regularly, especially those with flow cytometry data, there are 5 specific questions I ask when reading a manuscript.
If these questions are not adequately answered or explained in the paper, it raises red flags and ultimately leads to rejection.
1. What is the scientific hypothesis?
It all begins with the scientific hypothesis being tested. One of the critical parts of the paper is a clear statement of the reason for the study.
While it may seem ...Read More