Of Biomedical Molecular Biology, Cancer Research Institute Ghent (CRIG), Ghent University, Molecular and Cellular Oncology Lab, Inflammation Investigation Centre, VIB, Ghent, Belgium; 5Department of Biochemistry, Faculty of Medicine and Overall health Sciences, Ghent University, Ghent, Belgium; 6Institute for Transfusion Medicine, SIK1 Biological Activity University Hospital Essen, University of DuisburgEssen, Essen, CYP1 Storage & Stability Germany, Division of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden; 7Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Victoria, Australia; eight La Trobe Institute for Molecular Science; 9Department of Biochemistry Cell Biology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands; 10School of Pharmacy and Pharmaceutical Sciences and Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland; 11 Division of Animal Physiology and Immunology, TUM School of Life Sciences Weihenstephan, Technical University Munich, Munich, Germany; 12 Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, New York, USA; 13Laboratory of Lipid Metabolism and Cancer, Division of Oncology, LKI Leuven Cancer Institute, KU Leuven, Leuven, Belgium; 14 Institut Curie, PSL Study University, INSERM U932, Paris, France; 15 Institut Curie, PSL Investigation University, CNRS, UMR 144, Paris, France; 16 The Johns Hopkins University School of Medicine; 17Laboratory of Experimental Cancer Investigation, Division of Radiation Oncology and Experimental Cancer Investigation, Cancer Analysis Institute Ghent (CRIG), Ghent University, Ghent, BelgiumIntroduction: Extracellular vesicles (EVs) are critical intercellular communication cars for bioactive molecules with diagnostic and therapeutic relevance. The current growth of studies on EV effects in disease pathogenesis, tissue regeneration, and immunomodulation has led towards the application of numerous isolation and characterisation strategies poorly standardised and with scarcely comparable outcomes. Existing solutions for EV characterisation mainly rely on common biomarkers and physical options that don’t mirror the actual heterogeneity of vesicles. Raman spectroscopy is a label-free, fast, non-destructive, sensitive method that can turn into a beneficial tool for the biochemical characterisation and discrimination of EVs from several cell types. Methods: Human mesenchymal stromal cells from bone marrow and adipose tissue, and dermal fibroblasts were cultured for 72 h in serum no cost conditions. Ultracentrifuged vesicles obtained from conditioned media have been analysed by confocal Raman microspectroscopy with 532 nm laser sources within the spectral ranges 500800 cm-1 and 2600200 cm-1. Multivariate statistical evaluation (PCA-LDA) and classical least squares (CLS) fitting with reference lipid molecules (cholesterol, ceramide, phosphatidylcholine, phosphatidylethanolamine, phosphatidic acid and GM1) had been performed on recordings obtained on air-dried drops of EV suspensions. Benefits: When vesicles had been irradiated, Raman bands of nucleic acids, proteins, and lipids (cholesterol, phospholipids) have been visible inside the spectra providing a biochemical fingerprint of your deemed vesicles. CLS fitting permitted the calculation on the relative contribution of lipids to the recorded spectra. By Raman spectroscopy we can clearly distinguish vesicles originated by distinctive cell-types with great accuracy (around 93) because of biochemical attributes common of the.
