A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture

Traditionally, midbody remnants (MBRs) are isolated from cell culture medium using ultracentrifugation, which is expensive and time consuming. Here, we present a protocol for isolating MBRs or large extracellular vesicles (EVs) from mammalian cell culture using either 1.5% polyethylene glycol 6000 (...

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Autores principales: Park, Sungjin, Patel, Smit A., Torr, Elizabeth E., Dureke, Ashley-Grace N., McIntyre, Alina M., Skop, Ahna R.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2023
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10500451/
https://www.ncbi.nlm.nih.gov/pubmed/37690025
http://dx.doi.org/10.1016/j.xpro.2023.102562
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author Park, Sungjin
Patel, Smit A.
Torr, Elizabeth E.
Dureke, Ashley-Grace N.
McIntyre, Alina M.
Skop, Ahna R.
author_facet Park, Sungjin
Patel, Smit A.
Torr, Elizabeth E.
Dureke, Ashley-Grace N.
McIntyre, Alina M.
Skop, Ahna R.
author_sort Park, Sungjin
collection PubMed
description Traditionally, midbody remnants (MBRs) are isolated from cell culture medium using ultracentrifugation, which is expensive and time consuming. Here, we present a protocol for isolating MBRs or large extracellular vesicles (EVs) from mammalian cell culture using either 1.5% polyethylene glycol 6000 (PEG6000) or PEG5000-coated gold nanoparticles. We describe steps for growing cells, collecting media, and precipitating MBRs and EVs from cell culture medium. We then detail characterization of MBRs through immunofluorescent antibody staining and immunofluorescent imaging.
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spelling pubmed-105004512023-09-15 A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture Park, Sungjin Patel, Smit A. Torr, Elizabeth E. Dureke, Ashley-Grace N. McIntyre, Alina M. Skop, Ahna R. STAR Protoc Protocol Traditionally, midbody remnants (MBRs) are isolated from cell culture medium using ultracentrifugation, which is expensive and time consuming. Here, we present a protocol for isolating MBRs or large extracellular vesicles (EVs) from mammalian cell culture using either 1.5% polyethylene glycol 6000 (PEG6000) or PEG5000-coated gold nanoparticles. We describe steps for growing cells, collecting media, and precipitating MBRs and EVs from cell culture medium. We then detail characterization of MBRs through immunofluorescent antibody staining and immunofluorescent imaging. Elsevier 2023-09-09 /pmc/articles/PMC10500451/ /pubmed/37690025 http://dx.doi.org/10.1016/j.xpro.2023.102562 Text en © 2023 The Authors https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Protocol
Park, Sungjin
Patel, Smit A.
Torr, Elizabeth E.
Dureke, Ashley-Grace N.
McIntyre, Alina M.
Skop, Ahna R.
A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
title A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
title_full A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
title_fullStr A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
title_full_unstemmed A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
title_short A protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
title_sort protocol for isolating and imaging large extracellular vesicles or midbody remnants from mammalian cell culture
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10500451/
https://www.ncbi.nlm.nih.gov/pubmed/37690025
http://dx.doi.org/10.1016/j.xpro.2023.102562
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