Osomal markers was carried through FACS employing microspheres and MASPlex exosome kit. MASPlex kit simultaneously detects 37 exosome surface epitopes. Benefits: We set up a process for EV isolation from AF determined by subsequent dilution with PBS; 1st centrifugation at 10,000 g for 30 min at 4 , filtration via a 0.45 filter and ultracentrifugation at 100,000 g for 2 h in 4 . The averages EV concentration was 4.34011 particles/ml having a imply peak of 240.45 nm, measured by NTA. FACS evaluation showed presence of angiogenic markers VEGFR 1,2,3 and CD105, immunological markers HLA ABC, HLA DR, exosome certain markers CD81 and CD63 also CD133, which indicates kidney origin. By using the MASPlex kit, we setup a semiquantitative technique for detection of 37 diverse potential AF-EV surface markers in 1 sample simultaneously. We confirmed the heterogenic FSH Receptor Proteins Biological Activity traits of AF-EVs, like expression of immune program markers CD209, CD62P, CD11c, CD20 and endothelial markers CD146 and CD41b.Summary/Conclusion: The characterisation from the AFEVs with NTA and FACS demonstrates the composition and size as well as presence of markers of different origin including kidney, immune system and endothelium. The investigation of EV properties in healthier and diseased placenta could prove valuable inside the future as a diagnostic tool to understand and diagnose pregnancyassociated diseases. Funding: This perform was supported by the iPlacenta project founded by the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No.PF09.Evaluation of non-invasive biomarkers for monitoring functional status of endometrium Mattia Criscuolia, Gaia Papinia, Davide Zoccob, Alice PD-1 Proteins Formulation Luddic, Valentina Pavonec, Paola Piombonic and Natasa ZarovnidaExosomics Siena University of Siena, Siena, Italy; bExosomics Siena, Siena, USA; cUniversity of Siena, Siena, Italy; dExosomics, Siena, ItalyIntroduction: Endometrium is a complicated tissue with self-renewing properties, typically undergoing cyclic modifications regulated by ovarian steroids divided into proliferative and secretory phase. The transcriptomic profile of the endometrium is influenced by other endometrial cell sorts (glandular epithelial and stromal) in both physiological and pathological conditions. These cells have mutual paracrine effects partially mediated by EVs, and they grow inside a cycledependent manner. To assess the endometrium status, a number of invasive or expensive methods are at present employed, including immunohistochemistry (IHC) on tissue biopsy, cytology and imaging. Development of protocols for the isolation of EVs from novel biological sources is definitely an particularly attractive signifies to surrogate endometrial biopsies. These novel protocols may perhaps enable the identification and sensitive detection of distinct endometrial EV biomarkers for diagnostic solutions in reproductive medicine, endometriosis or cancer. Solutions: Samples: key endometrial cultures, urine from wholesome donors in secretory phase; Differential centrifugation, size exclusion chromatography (SEC),JOURNAL OF EXTRACELLULAR VESICLESimmunobeads for EV isolation; Nanoparticle Tracking Analysis (NTA), BCA assay, ELISA, HS Qubit, ddPCR, SPR, FACS for EVs and EV markers quantification and characterization. Results: We provide new evidence that urine is really a surrogate biofluid appropriate for the detection of endometrial EV biomarkers. Utilizing pre-selected antibody panels, we determine distinct endometrium EV binding antib.