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  • Comprehensive Characterization of Human Dendritic Cells and Monocytes using High-Dimensional Flow Cytometry

    Contains 3 Component(s), Includes Credits Includes a Live Web Event on 10/06/2021 at 12:00 PM (EDT)

    A CYTO U webinar presented by Florian Mair, PhD and Thomas Liechti, PhD Keywords: Dendritic cells, Monocytes, High-dimensional flow cytometry, Panel design, Unsupervised data analysis

    About the Presenter

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    Thomas Liechti, PhD
    Postdoctoral Researcher
    ImmunoTechnology Section
    Vaccine Research Center
    National Institutes of Health

    Thomas Liechti obtained his PhD in immunology and microbiology at the University of Zurich in 2017 and is currently a postdoctoral researcher in Mario Roederer’s group at the Vaccine Research Center of the National Institutes of Health in Bethesda (USA). His main interest is high-dimensional flow cytometry and human immunology. During his postdoctoral training, he established a 28-color flow cytometry sample processing and analysis pipeline to assess the contribution of genetic and environmental factors to human immune homeostasis in a cohort of more than 3000 individuals.  


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    Florian Mair, PhD
    Research Associate | Cytometry Specialist
    Fred Hutchinson Cancer Research Center

    Florian Mair graduated with a PhD from the University of Zurich, Switzerland, in 2014 and is currently working at the Fred Hutchinson Cancer Research Center in Seattle, USA, as a research associate. During the past decade, he has been involved extensively with different cytometry platforms (conventional, spectral and mass cytometry) as well as scRNA-seq techniques and developed an interest in applying novel analysis approaches for single cell data. He has been actively engaged in teaching flow cytometry courses, including systematic panel design and analysis of high-dimensional cytometry experiments. 

    Webinar Summary

    Human dendritic cells (DCs) and monocytes are critical components of the innate immune system and important for orchestrating adaptive immunity. In recent years high-dimensional single cell technologies, such as flow cytometry and single cell RNA-seq, dissected the heterogeneity of human DCs and monocytes more precisely, suggesting more robust markers to unequivocally define these subsets. Based on these results and our recently published phenotype report (Mair F. and Liechti T. Cytometry Part A, 2021), we will discuss strategies to better define and characterize human DCs and monocytes using a combination of traditional and recently discovered markers. We will show practical examples of how the combination of thorough panel design and unsupervised data analysis can help to dissect heterogenous immune populations. This webinar will be split into three parts:

    • A description about the phenotypic and functional characteristics of human DC and monocyte subsets with a focus on how the emergence of single-cell technologies improved our understanding of the heterogenous landscape of human phagocytes.
    • High-dimensional panel design for characterization of human DCs and monocytes with a more specific focus on marker selection and the incorporation of biological knowledge into this process.
    • An overview of how unsupervised data analysis approaches can improve the delineation of human DC and monocyte subsets.

    Learning Objectives

    • Learn about the phenotypic and functional characteristics of human DCs and monocytes.
    • Understand strategies for marker selection in high-dimensional flow cytometry panel design.
    • Learn how unsupervised data analysis can guide the analysis of human DCs and monocytes.

     Who Should Attend

    Scientist interested in studying human dendritic cells and monocytes using flow cytometry. In addition, flow cytometrists with a general interest in high-dimensional flow cytometry panel design.

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • CYTO Virtual Interactive 2021 Hooke Lecture

    Contains 3 Component(s), Includes Credits

    CYTO Virtual Interactive 2021 Hooke Lecture Presented by Sharon Lewin, AO, FRACP, FAHMS

    Speaker

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    Sharon Lewin, AO, FRACP, FAHMS
    Director, Peter Doherty Institute for Infection and Immunity

    Session Summary

    Leading infectious diseases expert, Professor Sharon Lewin, is the inaugural Director of the Doherty Institute. She is also a Professor of Medicine at The University of Melbourne and a National Health and Medical Research Council (NHMRC) Practitioner Fellow. As an infectious diseases physician and basic scientist, her laboratory focuses on basic, translational and clinical research aimed at finding a cure for HIV and understanding the interaction between HIV and hepatitis B virus. Her laboratory is funded by the NHMRC, the National Institutes of Health, The Wellcome Trust, the American Foundation for AIDS Research and multiple commercial partnerships. She is also the Chief Investigator of a NHMRC Centre of Research Excellence (CRE), The Australian Partnership for Preparedness Research on Infectious Diseases Emergencies (APPRISE) that aims to bring together Australia’s leading experts in clinical, laboratory and public health research to address the key components required for a rapid and effective emergency response to infectious diseases.

    Panelists

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    Adrian Smith, PhD
    Cytometry, Imaging and IT Manager
    Centenary Institute

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    Alastair J. Sloan; BSc, PhD, PGCert, FHEA, CBiol, FRSB
    FICD Head
    Melbourne Dental School

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    Grace Chojnowski
    Flow Cytometry and Imaging Facility Manager
    Queensland Institute of Medical Research

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • What/Why Should I Care About Deep, Image-Based Biophysical Single-Cell Analysis?

    Contains 3 Component(s), Includes Credits Recorded On: 05/21/2021

    A CYTO U Webinar presented by Kevin Tsia, PhD Keywords: Imaging flow cytometry, high-throughput screening, single-cell analysis, biophysical cytometry

    About the Presenter

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    Kevin Tsia, PhD
    Professor
    University of Hong Kong

    Kevin Tsia received his PhD degree in electrical engineering at the University of California Los Angeles (UCLA) in 2009. He is currently a professor in the Department of Electrical and Electronic Engineering and the program director of the Biomedical Engineering Program at the University of Hong Kong (HKU). His research interest covers a broad range of subject matters including ultra-fast optical imaging for imaging flow cytometry and cell-based assay high-speed in-vivo brain imaging computational approaches for single-cell analysis. In 2012, he received the Early Career Award by the HK Research Grants Council in Hong Kong. He also received the Outstanding Young Research Award in 2015 at HKU as well as the 14th Chinese Science and Technology Award for Young Scientists in 2016. He is currently the RGC Research Fellow. He holds four-granted and four-pending US patents on ultrafast optical imaging technologies. He is co-founder of a start-up company commercializing the high-speed microscopy technology for cancer screening and treatment monitoring applications.

    Webinar Summary

    It has long been recognized that the association between the molecular genetic landscape that instructs expression of proteins and macromolecules in single cells is intrinsically linked with their biophysical properties (e.g., cell morphology, size, mass, force etc.). A growing body of evidence shows that the label-free assessment of biophysical properties of cells is an effective (or even more accurate) descriptor of cellular heterogeneity, compared to the conventional fluorescence markers, at single-cell precision. Furthermore, how molecular signatures translate into the emergent cellular biophysical properties has not been fully understood. Only with the recently advanced techniques can we now start to investigate this link.

    This webinar will introduce how the synergism among single-cell imaging, microfluidics, and deep learning allows us to overcome the current limitations of single-cell biophysical phenotyping (in both instrumentation integration and new data analytic strategies). Specifically, a few high-throughput, deep-learning-powered imaging techniques will be described, as well as cytometry pipelines developed in our laboratory over the past few years. These platforms allow researchers to significantly scale the single-cell biophysical phenotyping throughput (beyond millions of cells) and enrich the phenotyping content by integrating with biochemical cell-based assay in a single-platform. Pushing the limit of biophysical phenotyping specificity and sensitivity, these techniques have been successfully employed to a number of biological research and clinical applications, including rare cancer cell detection in mouse blood, cancer cell sub-typing, targeted-drug sensitivity prediction, and so on.

    Learning Objectives

    • Recent advances in biophysical cytometry, especially label-free single-cell imaging, that enables studies of cellular heterogeneity at the levels of throughput, precision, specificity, and sensitivity that were once inconceivable.
    • Advanced techniques, involving synergism among microfluidics, imaging and deep learning, that allow us to investigate deeper the link between molecular signatures and the emergent cellular biophysical properties. 

    Who Should Attend

    Biomedical scientists and engineers working on developments of cytometry platforms, single-cell imaging technologies, and single-cell analysis. 

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • The Road to a High-Resolution 40-Color Flow Cytometry Immunophenotyping Panel

    Contains 3 Component(s), Includes Credits

    A CYTO U Webinar presented by Maria C. Jaimes, MD Keywords: High-dimensional flow cytometry, spectral flow cytometry, OMIP, Panel development

    About the Presenter

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    Maria C. Jaimes, PhD
    VP Technical Application Support
    Cytek Biosciences, Inc.

    Dr. Maria Jaimes earned her MD degree at the Universidad Javeriana in Colombia. Dr. Jaimes completed her postdoctoral training at Stanford University in the Department of Microbiology and Immunology. During her postdoc, she focused on characterizing the immune responses to both rotavirus and influenza viruses after natural infection and immunization. In 2005, Dr. Jaimes joined BD Biosciences, where she worked on different aspects of quality assurance and standardization of flow cytometry assays. In 2015, Dr. Jaimes joined Cytek Biosciences. She is part of the R&D team credited for developing the Aurora Full Spectrum Cytometer and has overseen the instrument characterization, verification, and development of multicolor applications. Besides her responsibilities within the R&D team, Dr. Jaimes leads the Technical Applications Support team worldwide.

    Webinar Summary

    This webinar will cover OMIP-069 (published in Cytometry Part A in August 2020), the first 40-color fluorescent panel using full spectrum flow cytometry to broadly phenotype much of the cellular composition of the human peripheral immune system. The panel in this OMIP has been thoroughly optimized to ensure high-quality data and well-resolved populations, enabling the description of most canonical subsets of T cells, B cells, NK cells, monocytes, and dendritic cells. Dr. Jaimes will present the journey of the technology, panel design, protocol development, data QC/QA, and data analysis which led to the successful achievement of this 40-color immune profiling panel. 

    Learning Objectives

    • Understand the concepts behind full spectrum profiling.
    • Learn the necessary steps and tools for good panel design.
    • Develop expertise on how to troubleshoot and optimize a multicolor panel.
    • Learn to recognize high quality vs. compromised data and the potential sources and mitigation of errors. 

    Who Should Attend

    SRL staff/directors, immunologists, CRO staff, and full spectrum flow cytometer users.

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • Evaluating Spectral Cytometry for Immune Profiling in Viral Disease

    Contains 3 Component(s), Includes Credits

    A CYTO U Webinar presented by Paula Niewold, PhD and Thomas Ashhurst, PhD Keywords: spectral cytometry, unmixing, compensation, conventional cytometry, autofluorescence

    About the Presenters

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    Paula Niewold, PhD
    Postdoctoral Researcher
    Department of Infectious Diseases
    Leiden University Medical Centre

    Dr. Paula Niewold is an immunologist currently working as a postdoctoral researcher at the Department of Infectious Diseases at the Leiden University Medical Centre. She is interested in host-pathogen interactions and how they impact the outcome of disease. She has studied these interactions in models of cerebral malaria, West Nile virus encephalitis, psoriasis, and tuberculosis using high-dimensional flow, mass, and imaging mass cytometry. She is an ISAC Marylou Ingram Scholar.

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    Thomas Ashhurst, PhD
    Immunologist and High-Dimensional Cytometry Specialist
    Sydney Cytometry Facility
    University of Sydney

    Dr. Thomas Ashhurst is an immunologist and high-dimensional cytometry specialist with the Sydney Cytometry Facility at the University of Sydney. He develops and applies a range of single-cell cytometry technologies and computational analysis tools to map dynamic immune responses over time, space, and disease. In particular, he applies these approaches to the study of immunology and infectious disease, including emerging pathogens such as COVID-19, Zika virus encephalitis, and West Nile virus encephalitis. He is an ISAC Marylou Ingram Scholar.

    Webinar Summary

    In conventional fluorescence cytometry, each fluorophore in a panel is measured in a target detector, through the use of wide band-pass optical filters. In contrast, spectral cytometry uses a large number of detectors with narrow band-pass filters to measure a fluorophore's signal across the spectrum, creating a more detailed fluorescent signature for each fluorophore. The spectral approach shows promise in adding flexibility to panel design and improving the measurement of fluorescent signal. However, few comparisons between conventional and spectral systems have been reported to date. Here we present our findings comparing conventional and spectral approaches to cytometry—including comparisons of compensation and unmixing—and evaluate the use of spectral cytometry for immune profiling in viral diseases.

    Learning Objectives

    • Gain an understanding of the essential differences between conventional and spectral approaches to cytometry.
    • Appreciate the differences between compensation and spectral unmixing.
    • Consider applications for spectral cytometry in the context of immunological studies.

    Who Should Attend

    Researchers and technical staff who utilize flow, spectral, or mass cytometry in their work.

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • Proliferation Monitoring, Lesson 1: Basics of the Cell Cycle

    Contains 4 Component(s), Includes Credits

    Online, self-paced course presented by Kylie Price, Katherine (Kathy) Muirhead, and Paul Wallace

    Lesson 1 of the proliferation course reviews the basic concepts of cell cycle biology, setting the stage for the more specific methods and applications discussed in Lessons 2 and 3. Topics discussed include:

    • Phases of the cell cycle and molecular events associated with each.
    • Cell cycle checkpoints and regulators (cyclins and cyclin dependent kinases).
    • The relationship between the cell cycle phase distribution of a sample and DNA content histogram.
    • The use of DNA content histograms in combination with metabolic labeling or cellular synchronization to determine kinetics of progression through the different cell cycle phases.

    Authors:

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    Kylie M. Price
    Hugh Green Cytometry Core Facility
    Malaghan Institute of Medical Research
    Wellington, New Zealand

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    Katharine A. (Kathy) Muirhead
    SciGro Inc., Midwest Office
    Middleton, Wisconsin, USA

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    Paul K. Wallace
    Department of Flow and Image Cytometry
    Roswell Park Cancer Institute
    Buffalo, New York, USA

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • Proliferation Monitoring, Lesson 2B: Data Analysis Strategies

    Contains 4 Component(s), Includes Credits

    Online, self-paced course presented by Kylie Price, Kathy Muirhead, and Paul Wallace

    This is the second of four modules of Lesson 2 of the Proliferation course. Module 2B provides an in depth discussion of DNA cell cycle data analysis strategies. The underlying concepts behind the characteristic cell cycle distribution are described followed by a description with examples of simple and complex approaches to modeling normal and abnormal DNA fluorescence data.

    The course concludes with a discussion of controls to include in proper experimental design and explains how to analyze synchronized cell populations as seen when cell cultures are drug treated. Anyone with a basic understanding of flow cytometry who is unfamiliar with (or wishes to review) how DNA flow cytometric data is analyzed is encouraged to attend.

    Authors

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    Kylie M. Price
    Hugh Green Cytometry Core Facility
    Malaghan Institute of Medical Research
    Wellington, New Zealand

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    Katharine A. (Kathy) Muirhead 
    SciGro, Inc., Midwest Office
    Middleton, Wisconsin, USA

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    Paul K. Wallace  
    Department of Flow and Image Cytometry
    Roswell Park Cancer Institute
    Buffalo, New York, USA

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • Proliferation Monitoring, Lesson 2C: Practical Considerations & Troubleshooting

    Contains 4 Component(s), Includes Credits

    Online, self-paced course presented by Kylie Price, Kathy Muirhead, and Paul Wallace

    This is the third of four modules in Lesson 2 of the proliferation course, which focuses on cell cycle analysis by flow cytometry. Module 2C: “Practical Considerations and Troubleshooting" examines how choices made during sample preparation, reagent selection, instrument setup, and data collection impact the reliability and quality of the resulting DNA histograms. Topics covered include instrument linearity, fixation effects, doublet discrimination techniques, and special considerations for DNA cell cycle analysis using vital DNA dyes and thymidine analogs.

    Authors

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    Kylie M. Price  
    Hugh Green Cytometry Core Facility
    Malaghan Institute of Medical Research
    Wellington, New Zealand

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    Katharine A. (Kathy) Muirhead 
    SciGro Inc., Midwest Office
    Middleton, Wisconsin, USA

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    Paul K. Wallace  
    Department of Flow and Image Cytometry
    Roswell Park Cancer Institute
    Buffalo, New York, USA


    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • Proliferation Monitoring, Lesson 2D: Applications

    Contains 4 Component(s), Includes Credits

    Online, self-paced course presented by Kylie Price, Kathy Muirhead, and Paul Wallace

    This is the final module in Lesson 2 of the proliferation course, which focuses on cell cycle analysis by flow cytometry. Module 2D “Applications" looks at how investigators have used the principles and methods discussed in Lessons 2A⁠–⁠2C to monitor cell cycle status and cell cycle progression in their research fields of interest. Topics covered include tumor diagnosis and prognosis, anti-tumor mechanisms of therapeutic agents, and regulation of growth versus quiescence or senescence in normal and neoplastic cells. 

    Authors

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    Kylie M. Price
    Hugh Green Cytometry Core Facility
    Malaghan Institute of Medical Research
    Wellington, New Zealand

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    Katharine A. (Kathy) Muirhead
    SciGro Inc., Midwest Office
    Middleton, Wisconsin, USA

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    Paul K. Wallace
    Department of Flow and Image Cytometry
    Roswell Park Cancer Institute
    Buffalo, New York, USA


    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information
  • Reproducibility Crisis and Antibody Validation of Flow Cytometry

    Contains 4 Component(s), Includes Credits Recorded On: 03/04/2021

    A CYTO U Webinar presented by Pablo Engel, PhD Keywords: antibodies, reagent validation, reproducibility

    About the Presenter

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    Pablo Engel, PhD
    Professor of Immunology
    University of Barcelona

    Pablo Engel obtained his MD and PhD from the University of Barcelona. After training as a postdoctoral fellow at the Dana-Farber Cancer Institute of Harvard Medical School, Pablo became an assistant professor in the Department of Immunology at Duke University School of Medicine. He is currently a professor of immunology and head of the Immunology Unit in the Department of Biomedical Sciences at the University of Barcelona. His research is focused on lymphocyte cell-surface molecules and their role in the regulation of immune responses. He is also an expert in the production and characterization of monoclonal antibodies. In addition to his research, Pablo is Secretary General of the European Federation of Immunological Societies (EFIS).

    Webinar Summary

    The presentation will highlight the current reproducibility crisis, specifically reflecting on the effect of poorly validated antibodies on the research. The main causes of antibody failure will also be reviewed, as well as a basic antibody validation protocol for flow cytometry. Several solutions will be discussed to solve the problem of reproducibility. 

    Learning Objectives

    • To understand the urgent need of antibody validation.
    • To gain knowledge about basic antibody validation protocols.

    Who Should Attend

    Scientists and technologist that use monoclonal antibodies of flow cytometry.

    CMLE Credit: 1.0

    • Register
      • Non-member - Free!
      • Full - Free!
      • Student - Free!
      • SRL Junior Staff - Free!
      • SRL Emerging Leader - Free!
      • Scholar - Free!
      • Emeritus - Free!
      • Life - Free!
      • ISAC Staff - Free!
      • Community Administrator - Free!
      • Student Non-Member - Free!
      • SRL Junior Staff Non-member - Free!
      • Innovator - Free!
    • More Information