Chromatography and its hyphenation to mass spectrometry for extracellular vesicle analysis

Gabriella Pocsfalvi, Christopher Stanly, Immacolata Fiume, Károly Vékey

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Extracellular vesicles (EVs), such as exosomes, microvesicles and apoptotic bodies are released by cells, both under physiological and pathological conditions. EVs can participate in a novel type of intercellular communication and deliver cargo of nucleic acids, proteins and lipids near or to distant host cells. EV research is proceeding at a fast pace; now they start to appear as promising therapeutic targets, diagnostic tools and drug delivery systems. Isolation and analysis of EVs are prerequisites for understanding their biological roles and for their clinical exploitation. In this process chromatography and mass spectrometry (MS)-based strategies are rapidly gaining importance; and are reviewed in the present communication.Isolation and purification of EVs is mostly performed by ultracentrifugation at present. Chromatography-based strategies are gaining ground, among which affinity and size exclusion chromatography (SEC) are particularly strong contenders. Their major advantages are the relative simplicity, robustness and throughput. Affinity chromatography has the added advantage of separating EV subtypes based on molecular recognition of EV surface motifs. SEC has the advantage that isolated EVs may retain their biological activity.EVs are typically isolated in small amounts, therefore high sensitivity is required for their analysis. Study of the molecular content of EVs (all compounds beside nucleic acids) is predominantly based on liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. The chromatographic separation is mostly performed by reverse phase, nanoscale, ultra high performance LC technique. The MS analysis relying typically on nano-electrospray ionization MS/MS provides high sensitivity, selectivity and resolution, so that thousand(s) of proteins can be detected/identified/quantified in a EV sample. Beside protein identification, quantitation and characterization of protein post-translational modifications (PTMs), like glycosylation and phosphorylation are becoming feasible and increasingly important. Along with conventional LC-MS/MS, other chromatographic approaches hyphenated to MS are gaining importance for EV characterization. Hydrophilic interaction LC is used to characterize PTMs; LC-inductively coupled plasma/MS to identify metal containing molecules; while gas chromatography-MS to analyze some lipids and metabolites.

Original languageEnglish
Pages (from-to)26-41
Number of pages16
JournalJournal of Chromatography A
Volume1439
DOIs
Publication statusPublished - Mar 25 2016

Fingerprint

Chromatography
Mass spectrometry
Mass Spectrometry
Tandem Mass Spectrometry
Size exclusion chromatography
Nucleic Acids
Post Translational Protein Processing
Proteins
Extracellular Vesicles
Gel Chromatography
Glycosylation
Lipids
Affinity chromatography
Inductively coupled plasma mass spectrometry
Electrospray ionization
Molecular recognition
Phosphorylation
Exosomes
Communication
Liquid chromatography

Keywords

  • Affinity chromatography
  • Extracellular vesicles (EVs)
  • Hyphenated techniques
  • Mass spectrometry (MS)
  • Purification
  • Size-exclusion chromatography (SEC)

ASJC Scopus subject areas

  • Analytical Chemistry
  • Organic Chemistry
  • Biochemistry

Cite this

Chromatography and its hyphenation to mass spectrometry for extracellular vesicle analysis. / Pocsfalvi, Gabriella; Stanly, Christopher; Fiume, Immacolata; Vékey, Károly.

In: Journal of Chromatography A, Vol. 1439, 25.03.2016, p. 26-41.

Research output: Contribution to journalArticle

Pocsfalvi, Gabriella ; Stanly, Christopher ; Fiume, Immacolata ; Vékey, Károly. / Chromatography and its hyphenation to mass spectrometry for extracellular vesicle analysis. In: Journal of Chromatography A. 2016 ; Vol. 1439. pp. 26-41.
@article{746eb3645a2b4387854ab34e6bd2bb81,
title = "Chromatography and its hyphenation to mass spectrometry for extracellular vesicle analysis",
abstract = "Extracellular vesicles (EVs), such as exosomes, microvesicles and apoptotic bodies are released by cells, both under physiological and pathological conditions. EVs can participate in a novel type of intercellular communication and deliver cargo of nucleic acids, proteins and lipids near or to distant host cells. EV research is proceeding at a fast pace; now they start to appear as promising therapeutic targets, diagnostic tools and drug delivery systems. Isolation and analysis of EVs are prerequisites for understanding their biological roles and for their clinical exploitation. In this process chromatography and mass spectrometry (MS)-based strategies are rapidly gaining importance; and are reviewed in the present communication.Isolation and purification of EVs is mostly performed by ultracentrifugation at present. Chromatography-based strategies are gaining ground, among which affinity and size exclusion chromatography (SEC) are particularly strong contenders. Their major advantages are the relative simplicity, robustness and throughput. Affinity chromatography has the added advantage of separating EV subtypes based on molecular recognition of EV surface motifs. SEC has the advantage that isolated EVs may retain their biological activity.EVs are typically isolated in small amounts, therefore high sensitivity is required for their analysis. Study of the molecular content of EVs (all compounds beside nucleic acids) is predominantly based on liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. The chromatographic separation is mostly performed by reverse phase, nanoscale, ultra high performance LC technique. The MS analysis relying typically on nano-electrospray ionization MS/MS provides high sensitivity, selectivity and resolution, so that thousand(s) of proteins can be detected/identified/quantified in a EV sample. Beside protein identification, quantitation and characterization of protein post-translational modifications (PTMs), like glycosylation and phosphorylation are becoming feasible and increasingly important. Along with conventional LC-MS/MS, other chromatographic approaches hyphenated to MS are gaining importance for EV characterization. Hydrophilic interaction LC is used to characterize PTMs; LC-inductively coupled plasma/MS to identify metal containing molecules; while gas chromatography-MS to analyze some lipids and metabolites.",
keywords = "Affinity chromatography, Extracellular vesicles (EVs), Hyphenated techniques, Mass spectrometry (MS), Purification, Size-exclusion chromatography (SEC)",
author = "Gabriella Pocsfalvi and Christopher Stanly and Immacolata Fiume and K{\'a}roly V{\'e}key",
year = "2016",
month = "3",
day = "25",
doi = "10.1016/j.chroma.2016.01.017",
language = "English",
volume = "1439",
pages = "26--41",
journal = "Journal of Chromatography",
issn = "0021-9673",
publisher = "Elsevier",

}

TY - JOUR

T1 - Chromatography and its hyphenation to mass spectrometry for extracellular vesicle analysis

AU - Pocsfalvi, Gabriella

AU - Stanly, Christopher

AU - Fiume, Immacolata

AU - Vékey, Károly

PY - 2016/3/25

Y1 - 2016/3/25

N2 - Extracellular vesicles (EVs), such as exosomes, microvesicles and apoptotic bodies are released by cells, both under physiological and pathological conditions. EVs can participate in a novel type of intercellular communication and deliver cargo of nucleic acids, proteins and lipids near or to distant host cells. EV research is proceeding at a fast pace; now they start to appear as promising therapeutic targets, diagnostic tools and drug delivery systems. Isolation and analysis of EVs are prerequisites for understanding their biological roles and for their clinical exploitation. In this process chromatography and mass spectrometry (MS)-based strategies are rapidly gaining importance; and are reviewed in the present communication.Isolation and purification of EVs is mostly performed by ultracentrifugation at present. Chromatography-based strategies are gaining ground, among which affinity and size exclusion chromatography (SEC) are particularly strong contenders. Their major advantages are the relative simplicity, robustness and throughput. Affinity chromatography has the added advantage of separating EV subtypes based on molecular recognition of EV surface motifs. SEC has the advantage that isolated EVs may retain their biological activity.EVs are typically isolated in small amounts, therefore high sensitivity is required for their analysis. Study of the molecular content of EVs (all compounds beside nucleic acids) is predominantly based on liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. The chromatographic separation is mostly performed by reverse phase, nanoscale, ultra high performance LC technique. The MS analysis relying typically on nano-electrospray ionization MS/MS provides high sensitivity, selectivity and resolution, so that thousand(s) of proteins can be detected/identified/quantified in a EV sample. Beside protein identification, quantitation and characterization of protein post-translational modifications (PTMs), like glycosylation and phosphorylation are becoming feasible and increasingly important. Along with conventional LC-MS/MS, other chromatographic approaches hyphenated to MS are gaining importance for EV characterization. Hydrophilic interaction LC is used to characterize PTMs; LC-inductively coupled plasma/MS to identify metal containing molecules; while gas chromatography-MS to analyze some lipids and metabolites.

AB - Extracellular vesicles (EVs), such as exosomes, microvesicles and apoptotic bodies are released by cells, both under physiological and pathological conditions. EVs can participate in a novel type of intercellular communication and deliver cargo of nucleic acids, proteins and lipids near or to distant host cells. EV research is proceeding at a fast pace; now they start to appear as promising therapeutic targets, diagnostic tools and drug delivery systems. Isolation and analysis of EVs are prerequisites for understanding their biological roles and for their clinical exploitation. In this process chromatography and mass spectrometry (MS)-based strategies are rapidly gaining importance; and are reviewed in the present communication.Isolation and purification of EVs is mostly performed by ultracentrifugation at present. Chromatography-based strategies are gaining ground, among which affinity and size exclusion chromatography (SEC) are particularly strong contenders. Their major advantages are the relative simplicity, robustness and throughput. Affinity chromatography has the added advantage of separating EV subtypes based on molecular recognition of EV surface motifs. SEC has the advantage that isolated EVs may retain their biological activity.EVs are typically isolated in small amounts, therefore high sensitivity is required for their analysis. Study of the molecular content of EVs (all compounds beside nucleic acids) is predominantly based on liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. The chromatographic separation is mostly performed by reverse phase, nanoscale, ultra high performance LC technique. The MS analysis relying typically on nano-electrospray ionization MS/MS provides high sensitivity, selectivity and resolution, so that thousand(s) of proteins can be detected/identified/quantified in a EV sample. Beside protein identification, quantitation and characterization of protein post-translational modifications (PTMs), like glycosylation and phosphorylation are becoming feasible and increasingly important. Along with conventional LC-MS/MS, other chromatographic approaches hyphenated to MS are gaining importance for EV characterization. Hydrophilic interaction LC is used to characterize PTMs; LC-inductively coupled plasma/MS to identify metal containing molecules; while gas chromatography-MS to analyze some lipids and metabolites.

KW - Affinity chromatography

KW - Extracellular vesicles (EVs)

KW - Hyphenated techniques

KW - Mass spectrometry (MS)

KW - Purification

KW - Size-exclusion chromatography (SEC)

UR - http://www.scopus.com/inward/record.url?scp=84960448631&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84960448631&partnerID=8YFLogxK

U2 - 10.1016/j.chroma.2016.01.017

DO - 10.1016/j.chroma.2016.01.017

M3 - Article

C2 - 26830636

AN - SCOPUS:84960448631

VL - 1439

SP - 26

EP - 41

JO - Journal of Chromatography

JF - Journal of Chromatography

SN - 0021-9673

ER -