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Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation

Recent years have brought great focus on the development of drug delivery systems based on extracellular vesicles (EVs). Considering the possible applications of EVs as drug carriers, the isolation process is a crucial step. To solve the problems involved in EV isolation, we developed and validated...

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Autores principales: Drożdż, Anna, Kamińska, Agnieszka, Surman, Magdalena, Gonet-Surówka, Agnieszka, Jach, Robert, Huras, Hubert, Przybyło, Małgorzata, Stępień, Ewa Łucja
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7558926/
https://www.ncbi.nlm.nih.gov/pubmed/32933147
http://dx.doi.org/10.3390/pharmaceutics12090872
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author Drożdż, Anna
Kamińska, Agnieszka
Surman, Magdalena
Gonet-Surówka, Agnieszka
Jach, Robert
Huras, Hubert
Przybyło, Małgorzata
Stępień, Ewa Łucja
author_facet Drożdż, Anna
Kamińska, Agnieszka
Surman, Magdalena
Gonet-Surówka, Agnieszka
Jach, Robert
Huras, Hubert
Przybyło, Małgorzata
Stępień, Ewa Łucja
author_sort Drożdż, Anna
collection PubMed
description Recent years have brought great focus on the development of drug delivery systems based on extracellular vesicles (EVs). Considering the possible applications of EVs as drug carriers, the isolation process is a crucial step. To solve the problems involved in EV isolation, we developed and validated a new EV isolation method—low-vacuum filtration (LVF)—and compared it with two commonly applied procedures—differential centrifugation (DC) and ultracentrifugation (UC). EVs isolated from endothelial cell culture media were characterized by (a) Transmission Electron Microscopy (TEM), (b) Nanoparticle Tracking Analysis (NTA), (c) Western blot and (d) Attenuated Total Reflection Fourier-Transform Infrared Spectroscopy (ATR-FTIR). Additionally, the membrane surface was imaged with Environmental Scanning Electron Microscopy (ESEM). We found that LVF was a reproducible and efficient method for EV isolation from conditioned media. Additionally, we observed a correlation between ATR-FTIR spectra quality and EV and protein concentration. ESEM imaging confirmed that the actual pore diameter was close to the values calculated theoretically. LVF is an easy, fast and inexpensive EV isolation method that allows for the isolation of both ectosomes and exosomes from high-volume sources with good repeatability. We believe that it could be an efficient alternative to commonly applied methods.
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spelling pubmed-75589262020-10-26 Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation Drożdż, Anna Kamińska, Agnieszka Surman, Magdalena Gonet-Surówka, Agnieszka Jach, Robert Huras, Hubert Przybyło, Małgorzata Stępień, Ewa Łucja Pharmaceutics Article Recent years have brought great focus on the development of drug delivery systems based on extracellular vesicles (EVs). Considering the possible applications of EVs as drug carriers, the isolation process is a crucial step. To solve the problems involved in EV isolation, we developed and validated a new EV isolation method—low-vacuum filtration (LVF)—and compared it with two commonly applied procedures—differential centrifugation (DC) and ultracentrifugation (UC). EVs isolated from endothelial cell culture media were characterized by (a) Transmission Electron Microscopy (TEM), (b) Nanoparticle Tracking Analysis (NTA), (c) Western blot and (d) Attenuated Total Reflection Fourier-Transform Infrared Spectroscopy (ATR-FTIR). Additionally, the membrane surface was imaged with Environmental Scanning Electron Microscopy (ESEM). We found that LVF was a reproducible and efficient method for EV isolation from conditioned media. Additionally, we observed a correlation between ATR-FTIR spectra quality and EV and protein concentration. ESEM imaging confirmed that the actual pore diameter was close to the values calculated theoretically. LVF is an easy, fast and inexpensive EV isolation method that allows for the isolation of both ectosomes and exosomes from high-volume sources with good repeatability. We believe that it could be an efficient alternative to commonly applied methods. MDPI 2020-09-13 /pmc/articles/PMC7558926/ /pubmed/32933147 http://dx.doi.org/10.3390/pharmaceutics12090872 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Drożdż, Anna
Kamińska, Agnieszka
Surman, Magdalena
Gonet-Surówka, Agnieszka
Jach, Robert
Huras, Hubert
Przybyło, Małgorzata
Stępień, Ewa Łucja
Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation
title Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation
title_full Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation
title_fullStr Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation
title_full_unstemmed Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation
title_short Low-Vacuum Filtration as an Alternative Extracellular Vesicle Concentration Method: A Comparison with Ultracentrifugation and Differential Centrifugation
title_sort low-vacuum filtration as an alternative extracellular vesicle concentration method: a comparison with ultracentrifugation and differential centrifugation
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7558926/
https://www.ncbi.nlm.nih.gov/pubmed/32933147
http://dx.doi.org/10.3390/pharmaceutics12090872
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