Abstract
Membrane vesicles are produced by all Gram-negative and Gram-positive bacteria investigated so far. Membrane vesicles are spherical bilayers of phospholipids released by the bacteria to their surrounding environment and whose average size is comprised between 20 and 300 nm. The purification of these vesicles is often a challenge, as the yield and purity are often crucial for further analyses or use. In this chapter, we describe the most used method to isolate membrane vesicles from culture supernatant of Streptococcus pneumoniae and Klebsiella pneumoniae using ultracentrifugation followed by a density gradient method.
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References
Alloing G, Granadel C, Morrison DA, Claverys JP (1996) Competence pheromone, oligopeptide permease, and induction of competence in Streptococcus pneumoniae. Mol Microbiol 21:471–478. https://doi.org/10.1111/j.1365-2958.1996.tb02556.x
Avila-Calderón ED, Ruiz-Palma M del S, Aguilera-Arreola MaG, Velázquez-Guadarrama N, Ruiz EA, Gomez-Lunar Z, Witonsky S, Contreras-Rodríguez A (2021) Outer membrane vesicles of gram-negative bacteria: an outlook on biogenesis. Front Microbiol 12:557902. https://doi.org/10.3389/fmicb.2021.557902
Briaud P, Carroll RK (2020) Extracellular vesicle biogenesis and functions in gram-positive bacteria. Infect Immun 88:e00433–e00420. https://doi.org/10.1128/IAI.00433-20
Ford T, Graham J, Rickwood D (1994) Iodixanol: a nonionic iso-osmotic centrifugation medium for the formation of self-generated gradients. Anal Biochem 220:360–366. https://doi.org/10.1006/abio.1994.1350
Gorringe A, Halliwell D, Matheson M, Reddin K, Finney M, Hudson M (2005) The development of a meningococcal disease vaccine based on Neisseria lactamica outer membrane vesicles. Vaccine 23:2210–2213. https://doi.org/10.1016/j.vaccine.2005.01.055
Klimentová J, Stulík J (2015) Methods of isolation and purification of outer membrane vesicles from gram-negative bacteria. Microbiol Res 170:1–9. https://doi.org/10.1016/j.micres.2014.09.006
Kulp A, Kuehn MJ (2010) Biological functions and biogenesis of secreted bacterial outer membrane vesicles. Annu Rev Microbiol 64:163–184. https://doi.org/10.1146/annurev.micro.091208.073413
McMillan HM, Kuehn MJ (2021) The extracellular vesicle generation paradox: a bacterial point of view. EMBO J 40:e108174. https://doi.org/10.15252/embj.2021108174
Narciso AR, Iovino F, Thorsdottir S, Mellroth P, Codemo M, Spoerry C, Righetti F, Muschiol S, Normark S, Nannapaneni P, Henriques-Normark B (2022) Membrane particles evoke a serotype-independent cross-protection against pneumococcal infection that is dependent on the conserved lipoproteins MalX and PrsA. Proc Natl Acad Sci U S A 119:e2122386119. https://doi.org/10.1073/pnas.2122386119
Schwechheimer C, Kuehn MJ (2015) Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions. Nat Rev Microbiol 13:605–619. https://doi.org/10.1038/nrmicro3525
Toyofuku M, Nomura N, Eberl L (2019) Types and origins of bacterial membrane vesicles. Nat Rev Microbiol 17:13–24. https://doi.org/10.1038/s41579-018-0112-2
Veerman RE, Teeuwen L, Czarnewski P, Güclüler Akpinar G, Sandberg A, Cao X, Pernemalm M, Orre LM, Gabrielsson S, Eldh M (2021) Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin. J Extracell Vesicles 10:e12128. https://doi.org/10.1002/jev2.12128
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Narciso, A.R., Aschtgen, MS. (2023). Strategies to Isolate Extracellular Vesicles from Gram-Negative and Gram-Positive Bacteria. In: Nordenfelt, P., Collin, M. (eds) Bacterial Pathogenesis. Methods in Molecular Biology, vol 2674. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3243-7_4
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DOI: https://doi.org/10.1007/978-1-0716-3243-7_4
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