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Expert Cytometry Webinars

SCYM(ASCP) Review: Section 2: Panel/Experimental Design

$19.99

The Certification in Cytometry credential, originally created by ISAC and ICCS, with a grant from the Wallace H Coulter foundation was designed to provide a tool to allow cytometrists to demonstrate a core set of knowledge necessary to design, execute, and interpret cytometry experiments. In 2017, ISAC, ICCS, and ASCP introduced the SPecialist in Cytometry (SCYM) credential to replace both the CCy and QCYM credentials.  The SCYM exam is composed of 100 questions, that are broken down into five major categories covering all aspects of the process, and including more clinically relevant questions for clinical cytometrists. The content outline and breakdown of the sections can be downloaded here. The eligibility requirements are found here This is the second in a series of lectures designed to focus on specific sections of the exam and help attendees acquire the knowledge that they will need to pass the exam. This lecture will focus on the Panel/Experimental Design section, which makes up between 25-30% of the exam. The specific topics to be covered are based on the SCYM exam outline, as listed below: Sample 

  • Sample source (e.g., beads, blood, bone marrow, solid tissue, body fluids, subcellular components, cultured cells, microorganisms, plants, whole organisms)
  1. Sample integrity (e.g., collection, handling, storage viability)
  2. Sample preparation and staining (e.g., disaggregation, lysing agents, aggregates, filtering, fixation, permeabilization)
  • Cell enrichment (e.g., cell sorting, density gradient isolation, magnetic beads)
Assay Development 
  • Target (e.g., cell type, subcellular location, molecule) 
  • Sample state for functional studies (e.g., activated, resting, proliferating) 
  • Probe types (e.g., antibodies, viability/DNA dyes, physiological, tracking dyes, fluorescent proteins)
  1. Fluorochrome selection (e.g., antigen density, protein coexpression, optimal combination, photostability, F/P ratio, quenching, signal to noise)
  1. Spectral overlap and compensation 
  2. Assay controls (e.g., fluorescence minus one (FMO), autofluorescence, biological systems control, background measurement controls)
  1. Assay optimization (e.g., appropriate use of limited sample, frequency of target, cell concentration, kinetics, scalability, blocking,
At the end of this webinar, you will have an idea of what topics will be on the SCYM exam and the type of material they need to be prepared to answer questions on. A brief quiz will be part of the lecture to allow attendees to see sample questions that may be asked on the exam.  Join Dr. Tim Bushnell, who helped develop the CCy content outline, as he reviews these important topics. 

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SCYM(ASCP) Review: Section 1: Instrumentation

$19.99

The Certification in Cytometry credential, originally created by ISAC and ICCS, with a grant from the Wallace H Coulter foundation was designed to provide a tool to allow cytometrists to demonstrate a core set of knowledge necessary to design, execute and interpret cytometry experiments. In 2017, ISAC, ICCS and ASCP introduced the SPecialist in Cytometry (SCYM) credential to replace both the CCy and QCYM credentials.  The SCYM exam is composed of 100 questions, that are broken down into five major categories covering all aspects of the process, and including more clinically relevant questions for clinical cytometrists. The content outline and breakdown of the sections can be downloaded here. The eligibility requirements are found here This is the first in a series of lectures designed to focus on specific sections of the exam, to help attendees prepare for the knowledge that they will need to help pass the exam. This lecture will focus on the instrumentation section, which makes up between 15-20% of the exam. The specific topics to be covered are based on the SCYM exam outline, as listed below:

  1. Fluidics 
  2. Hydrodynamic focusing and properties of sheath fluids 
  3. Generation of differential pressures (e.g., syringe pump, pressure based) 
  4. Optics 
  5. Optical filters (e.g., long pass, band pass, short pass, dichroics, neutral density, polarizing) 
  6. Light source (e.g., laser type, laser line, arc lamp, led) 
  7. Lenses (e.g., beam shape, collecting, focusing, objective) 
  8. Optical pathway (e.g., transmission, reflection, interrogation point, collinear, spatial
   separation, light scatter) 
  1. Electronics 
  2. Amplifiers (e.g., linear, logarithmic) 
  3. Detectors (e.g., photomultiplier tube, photodiode, CCD camera, avalanche photon detector) 
  4. Digital vs. analog systems 
  5. Noise 
  6. Pulse measurement (e.g., time delay, window extension, area, width, Coulter impedance) 
  7. Threshold/discriminator 

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Antibody Validation To Promote Reproducibility

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Antibodies are one of the most important reagents in flow cytometry experiments.  So when Bradbury and Plückthun published a commentary in Nature, where they estimated about 50% of the spending on ‘... protein binding reagents...’, was wasted because of ‘...poorly characterized and ill-defined antibodies...’ it served as a wakeup call to researchers to better understand both the causes of antibody failure and methods that could be used to optimize and validate their reagents.  There are many different factors that can impact the quality of a given antibody, and even the same clone from different vendors may perform differently.  

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Imaging Mass Cytometry

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Mass Cytometry (MC), as implemented by Fluidigm in the CyTOF, uses the principles of traditional flow cytometry (FFC) with one big difference.  In MC the reporter is a metal ion that has been tagged to the antibody rather than a fluorochrome.  Additionally, MC uses a time of flight detection system that is tuned to the mass range of the metals.  This technology allowed for the expansion into high dimensional analysis with 40+ parameters. By adding a laser to ablate a sample, releasing the bound ions that are taken into a TOF system, it became possible to develop complex images of the localization of the target antibodies, providing an in-depth visualization of the complex microenvironmental relationships present in the sample.  This has also been commercialized by Fluidigm in their Hyperion Imaging system, which can readily be added onto their Helios suspension system. This webinar is sponsored by Fluidigm. 

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5 Key Points In Flow Cytometry Data Analysis

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After the blood, sweat, and tears have been spilled, the experimental design optimization and validated, subjects consented, samples stained and the data has been collected, the work to interpret the data and reveal the secrets of the biology they are studying. Whether the work is discovery-driven or hypothesis-driven, there are many key factors that need to be understood and implemented to ensure the data is properly analyzed and that the analysis criteria can be communicated to the scientific community.  This is a critical factor to improve the reproducibility of the results.  This webinar will discuss five key points in flow cytometry data analysis and how to implement these points in your data analysis.

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