Proceedings of 7 th Socratic Lectures 2022 111 of 172 Scientific contribution/Original research Morphological Parameters of Erythrocyte Extracellular Vesicles at Hypoosmotic and Isoosmotic Conditions Jozelj M 1,2 , Košir T 1,2 , Božič D 1,2 , Hočevar M 3 , Pajnič M 2 , Iglič A 1,4 , Jeran M 1,2,& , Kralj-Iglič V 2, * 1. University of Ljubljana, Faculty of Electrical Engineering, Laboratory of Physics, Ljubljana, Slovenia 2. University of Ljubljana, Faculty of Health Sciences, Laboratory of Clinical Biophysics, Ljubljana, Slovenia 3. Department of Physics and Chemistry of Materials, Institute of Metals and Technology, Ljubljana, Slovenia 4. University of Ljubljana, Faculty of Medicine, Laboratory of Clinical Biophysics, Ljubljana, Slovenia * Correspondence: Veronika Kralj-Iglič; veronika.kralj-iglic@fe.uni-lj.si & Present address: “Jožef Stefan” Institute, Department of Inorganic Chemistry and Technology, Ljubljana, Slovenia JoM & KT contributed equally to this work. JeM and KIV contributed equally to this work. Abstract: Extracellular vesicles (EVs) are membrane-enclosed structures of nanometer dimensions. They are formed by cells and can be found in isolates from body fluids. It is indicated that they play important role in intercellular communication in health and disease. In this work we observed the morpholoical parameters of EVs isolated by differential centrifugation from washed erythrocytes, in which vesiculation was accelerated by addition of detergent (sodium dodecyl sulfate). Aliquots of the isolate were suspended medium of two different osmolarities. Isolates were imaged with a scan- ning electron microscope (SEM) and the images were analyzed by using the contours of EVs from which the volume V, surface area S, and relative volume v = (36S 3 /V 2 ) 1/2 were assessed by a geomet- rical model. EVs were considered as spheres or spheroids. The isolates were rich in erythrocyte EVs so that representative sets (86 vesicles at osmolarity of 50 mOsmol/L and 109 vesicles at osmolarity of 300 mOsmol/L) could be outlined. The EVs at osmolarity of 50 mOsmol/L had shapes close to a sphere, while at osmolarity of 300 mOsmol/L they had elongated shapes. The shapes were approxi- mated by prolate spheroids. The average volume/surface area of EVs at osmolarity 50 mOsmol/L were 3.18×10 5 nm 3 /2.20×10 4 nm 2, and at osmolarity 300 mOsmol/L they were 4.3×10 5 nm 3 /2.79×10 4 nm 2 . The respective differences in favor of the values at 300 mOsmol/L were statistically significant and of sufficient power. The relative volume of EVs at 50 mOsmol/L and 300 mOsmol/L were 1 and 0.96, respectively. While the difference in v suggests that, similarly to erythrocytes, water enters EVs in order to attain the Donnan equilibrium, the differences in V and A suggest that selective popping of (larger) EVs in the hypoosmolar sample took place. Keywords: Extracellular vesicles; Erythrocytes; Osmolarity; Differential centrifugation; Scanning electron microscopy (SEM); Morphology; Interdisciplinary connection; Medicine Citation: Jozelj M, Košir T, Božič D, Hočevar M, Pajnič M, Iglič A, Jeran M, Kralj-Iglič V. Using a physical approach to study mor- phological properties erythrocyte extracellular vesicles. Proceedings of Socratic Lectures. 2022; 7: 111- 115. https://doi.org/10.55295/PSL.2022. D16 Publisher’s Note: UL ZF stays neutral with regard to jurisdic- tional claims in published maps and institutional affiliations. Copyright: © 2022 by the authors. Submitted for possible open access publication under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/lice nses/by/4.0/).