APR 28, 2020 10:00 AM PDT

Measuring Single EVs and their Cargo: Sensitive and Specific Vesicle Flow Cytometry (vFCTM)

Speaker
  • Professor at the Scintillon Institute, a non-profit research institute and past director of the National Flow Cytometry Resource at Los Alamos
    Biography
      John Nolan, PhD is a professor at the Scintillon Institute, a non-profit research institute and past director of the National Flow Cytometry Resource at Los Alamos. He served as president of the International Society for Advancement of Cytometry (ISAC) from 2012-2014 and is a fellow of the American Institute of Medical and Biological Engineering. He has been working at the interface between technology and biology for more than twenty years to create and apply new tools for quantitative and multiplexed cell and molecular analysis and is recognized as a world leader in cytometry technology development. Dr. Nolan's recent work focuses on development of standardized flow cytometry-based methods and reagents for analysis of extracellular vesicles (EVs) to enable quantitative, reproducible EV characterization.

    Abstract
    DATE:  April 28, 2020
    TIME:  10:00am PT
     
    All cells release extracellular vesicles (EVs) that can carry molecular cargo to other cells to affect their function. EVs are promising as potential biomarkers, diagnostics and therapeutics, however understanding EV origins, compositional diversity, and biological effects is limited by our ability to measure them. EVs are heterogeneous, small and difficult to measure. Conventional biochemical and molecular methods (Western blot, ELISA, PCR) that report the total signal from target in a sample cannot resolve the identity of specific EVs, while single particle approaches (NTA, RPS, AFM) lack specificity and the ability to measure EV molecular cargo. Flow cytometry is attractive as a single vesicle measurement platform, but conventional instruments and assays lack the sensitivity and specificity to resolve EVs from other small particles and background, leading to artifacts and irreproducible, uninterpretable results. 

    A new generation of flow cytometers with high sensitivity photodetectors, together with the use of appropriate calibration protocols, has enabled the development of assays with the sensitivity and specificity to quantitatively and reproducibly measure individual EVs.  Vesicle Flow Cytometry (vFCTM, Cellarcus Biosciences), uses a membrane-selective dye to detect and size individual EVs together with optimized and validated antibodies and staining protocols to measure EV cargo. In this webinar, I will:
     
    1) Review the aims and approaches to EV analysis, including challenges for single EV measurement
    2) Demonstrate vFCTM using the Beckman Coulter CytoFLEX to count, size, and measure EV cargo
    3) Present data on the diversity of EV surface cargo expression, including tetraspanins, integrins, and tumor-associated markers, on individual EVs, and discuss the implications for understanding EV biology
     
    Webinars will be available for unlimited on-demand viewing after live event.
     
    LabRoots is approved as a provider of continuing education programs in the clinical laboratory sciences by the ASCLS P.A.C.E. ® Program. By attending this webinar, you can earn 1 Continuing Education credit once you have viewed the webinar in its entirety.

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