Used approach for characterization of extracellular vesicles (EVs). However, the applicability of flow cytometry has been somewhat limited resulting from the inability of conventional flow cytometers (FCM) to detect smaller sized EVs and discriminate between Complement Receptor 2 Proteins supplier Single events and so-called swarms of EVs. To overcome these challenges, recent advances in flow cytometry have led for the development of FCMs dedicated to the analysis of little particles (spFCM). Hence, the aim of this study should be to benchmark a novel FCM platform against a conventional FCM with regard to sensitivity, resolution and reproducibility in characterizing EVs directly in plasma. Techniques: Flow cytometry is performed on FACSAria III high-speed cell sorter (BD) and Apogee A60 Micro-PLUS (Apogee Flow Systems)Background: The heterogeneity of extracellular vesicles (EVs) needs new tools to characterize subpopulations and elucidate the effects and mechanisms by which they shape cellular processes. Not too long ago, important progress has been achieved in flow cytometry and fluorescence microscopy for high-throughput analysis of high-abundance markers in single EVs but none have yet been validated for single proteins on single vesicles. Here, we determine exosome-like extracellular vesicle (ELEV) subpopulations from breast cancer cell lines enriched on nanoarrays with single-ELEV resolution and single-molecule sensitivity. Procedures: A nanoarray of anti-mouse IgGs was printed onto a glass slide employing lift-off nanocontact printing, and the surface was passivated before incubation with mouse monoclonal capture antibodies. The nanoarray consists of 100 nm capture spots spaced 2 m apart that capture single ELEVs by virtue of their small size. ELEV samples, purified from cell supernatant applying size exclusion columns, have been incubated on the nanoarray overnight and detected making use of fluorescently tagged detection antibodies. Benefits: Single ELEV capture was demonstrated around the nanoarray utilizing AFM correlated with fluorescence microscopy. ELEVs could by detected with a single antibody as shown by single molecule photobleaching traces. Known exosome markers, integrins and basic cancer markers had been probed on exosomes derived from breast cancer cell lines, defining initial subpopulations. Summary/Conclusion: The heterogeneity of EVs calls for solutions that could measure single vesicles to enable for an accurate description of vesicle composition. Together with the nanoarray’s ability to enrich single ELEVs of interest in a high-throughput manner, ELEV subpopulations with one of a kind co-expression patterns can now be studied for their distinct effects. Funding: This study was funded by Genome Canada Disruptive Innovation in Genomics and NSERC.ISEV 2018 abstract bookPS09.Immunophenotyping extracellular vesicles by flow cytometry utilizing CCD-based imaging technology Sherree L. Friend; Haley R. Pugsley; Bryan Davidson; Phil Morrissey Merck KGaA/MilliporeSigma, Seattle, USABackground: Extracellular vesicles are membrane derived structures that include exosomes, microvesicles and apoptotic bodies. The value of extracellular vesicles as essential mediators of intercellular communication just isn’t effectively SARS-CoV-2 RNA Dependent RNA Polymerase Proteins manufacturer understood. Exosomes have already been shown to transfer molecules in between cells, potentially transmitting signals. Exosomes are released beneath regular physiological situations; however, they’re also believed to serve as mediators in the pathogenesis of neurological, vascular, haematological and autoimmune diseases at the same time as cancer. Quantifying and characterizi.