Isolation of extracellular membranous vesicles for proteomic analysis

Rommel A. Mathias, Justin W. Lim, Hong Ji, Richard J. Simpson

Research output: Chapter in Book/Report/Conference proceedingChapter (Book)Otherpeer-review

37 Citations (Scopus)


Membranous vesicles are constitutively released by a multitude of cell types. Following fusion of multivesicular bodies with the plasma membrane, endocytic vesicles, 30-90 nm in size termed exosomes are released extracellularly. Whilst several groups have reported the presence of exosomes in cell-culture conditioned medium, their biological and physiological functions still remain unclear. In addition, exosomes have been detected in body fluids associated with disease, further demonstrating their potential as diagnostic biomarkers. This protocol employs size filtration followed by ultracentrifugation to isolate and purify exosomes from the colon carcinoma cell line LIM 1215. Morphological visualisation and characterisation is based on electron microscopy and western blotting, whilst protein identification is achieved using a combination of 1D SDS-PAGE and LC-MS/MS.

Original languageEnglish
Title of host publicationMembrane Proteomics
Subtitle of host publicationMethods and Protocols
EditorsMatthew J. Peirce, Robin Wait
Place of PublicationTotowa, NJ USA
PublisherHumana Press
Number of pages16
ISBN (Print)9781603273091
Publication statusPublished - 2009
Externally publishedYes

Publication series

NameMethods in Molecular Biology
PublisherHumana Press
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

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