20.500.12556/RUL-137627 Extracellular vesicles from human cerebrospinal fluid are effectively separated by Sepharose CL-6B—comparison of four gravity-flow size exclusion chromatography methods Extracellular vesicles (EVs) are a versatile group of cell-secreted membranous nanoparticles present in body fluids. They have an exceptional diagnostic potential due to their molecular content matching the originating cells and accessibility from body fluids. However, methods for EV isolation are still in development, with size exclusion chromatography (SEC) emerging as a preferred method. Here we compared four types of SEC to isolate EVs from the CSF of patients with severe traumatic brain injury. A pool of nine CSF samples was separated by SEC columns packed with Sepharose CL-6B, Sephacryl S-400 or Superose 6PG and a ready-to-use qEV10/70 nm column. A total of 46 fractions were collected and analysed by slot-blot followed by Ponceau staining. Immunodetection was performed for albumin, EV markers CD9, CD81, and lipoprotein markers ApoE and ApoAI. The size and concentration of nanoparticles in fractions were determined by tunable resistive pulse sensing and EVs were visualised by transmission electron microscopy. We show that all four SEC techniques enabled separation of CSF into nanoparticle- and free protein-enriched fractions. Sepharose CL-6B resulted in a significantly higher number of separated EVs while lipoproteins were eluted together with free proteins. Our data indicate that Sepharose CL-6B is suitable for isolation of EVs from CSF and their separation from lipoproteins. extracellular vesicles cerebrospinal fluid traumatic brain injury size exclusion chromatography CD9 protein CD81 protein apolipoproteins zunajcelični vezikli cerebrospinalna tekočina travmatska poškodba možganov velikostno izločitvena kromatografija protein CD9 protein CD81 apolipoproteini true false true Angleški jezik Slovenski jezik Članek v reviji 2022-06-23 14:51:48 2022-06-23 14:51:50 2023-02-03 11:13:08 0000-00-00 00:00:00 2022 0 0 15 str. iss. 4, art. 785 Vol. 10 Apr. 2022 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 2227-9059 10.3390/biomedicines10040785 119230211 biomedicines-10-00785-v2_(1).pdf biomedicines-10-00785-v2_(1).pdf 1 181E55D03396290F1184333E8DF910BE 21c5fcc56aad3d8cde2888e655d17891c558fb54d8800df470adce410cc6b562 8a1ab6d4-f2f4-11ec-8aca-00155dcfd717 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=157654 https://www.mdpi.com/2227-9059/10/4/785 0 92ce99e3-f2f4-11ec-8aca-00155dcfd717 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=157655 Medicinska fakulteta 0 0 0