The aim of this study was to investigate cell source-dependent nucleic acids repertoire of diverse subpopulations of plasma extracellular vesicles (EVs). Blood plasma from nine healthy volunteers was used for the analysis. Samples of EVs were obtained by differential centrifugation of plasma. The application of high-sensitivity fluorescence-activated vesicles sorting (hs-FAVS) using fluorophore-conjugated anti-CD41-FITC (Fluorescein isothiocyanate) and anti-CD235a-PE antibodies allowed the isolation of three subpopulations of EVs, namely CD41+ CD235a-, CD41-CD235a+ and CD41-CD235a dim. The high purity (>97%) of the sorted subpopulations was verified by highsensitivity flow cytometry. Presence of nanosized objects in sorted samples was confirmed by combination of low-voltage scanning electron microscopy and dynamic light scattering. The amount of material in sorted samples was enough to perform Quantitative polymerase chain reaction (qPCR)-based nucleic acid quantification. The most prominent differences in the nucleic acid repertoire were noted between CD41+ CD235-vs. CD41-CD235a+ vesicles: the former contained significantly (p = 0.004) higher amount of mitochondrial DNA, and platelet enriched miR-21-5p (4-fold), miR-223-3p (38-fold) and miR-199a-3p (187-fold), but lower amount of erythrocyte enriched miR-451a (90-fold). CD41-CD235a+ and CD41-CD235a dim vesicles differed in levels of miR-451a (p = 0.016) and miR-21-5p (p = 0.031). Nuclear DNA was below the limit of detection in all EV subpopulations. The hs-FCM-based determination of the number of sorted EVs allowed the calculation of per single-event miRNA concentrations. It was demonstrated that the most abundant marker in CD41+ CD235a-subpopulation was miR-223-3p, reaching 38.2 molecules per event. In the CD41-CD235+ subpopulation, the most abundant marker was miR-451a, reaching 24.7 molecules per event. Taken together, our findings indicate that erythrocyte-and platelet-derived EVs carry different repertoires of nucleic acids, which were similar to the composition of their cellular sources.

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(UiO) in 2015. There I have met many fantastic people whose openness, enthusiasm, and disposition toward risky investments made this work possible. I will first greatly acknowledge Professor Jarle Vaage for inviting me to UiO and being my permanent co-supervisor. Thank you for invaluable help and for sharing your realistic optimism during all these exciting and challenging years. I am especially thankful for introducing me to Professor Guro Valen. Professor Guro Valen was a bright and inspiring supervisor under whose guidance we charted the blueprint of this work. She passed away in September 2014. I see no better way to express gratitude to her than being an active researcher. Professor Kåre-Olav Stensløkken has navigated me through the most challenging part of Ph.D., the final one. Thank you for the wise combination of understanding, proactive motivation, and especially for your patience! The triumvirate of Guro, Jarle, and Kåre-Olav fostered a unique working environmentour research group was always international, open, inclusive, and creative. It is my pleasure to name my colleagues from whom I have learned a lot: Arkady Rutkovsky, Lars Henrik Mariero, Marte Bliksøen, Apple Lei, and Fred Haugen were those with whom I have worked most with. Many thanks to doctor Rutkovsky for informal and all-purpose guidance during my first years in Norway. My special thanks to Torun Flatebø for numerous Westerns and PCRs you run for me. I would also like to give a tribute to all my previous colleagues: