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Table 2 Summary of common EV isolation methods

From: Basics and applications of tumor-derived extracellular vesicles

Method

Description

Example

Yield

Purity/quality

Equipment/Cost

Approximate processing time

Commercial products

Reference

Ultracentrifugation-based methods

Size- or density-dependent isolation

1. Differential centrifugation/Ultracentrifugation 2. Sucrose gradient ultracentrifugation

High

Protein aggregates and particulates contaminations

High equipment cost

2.5 – 48 h

No commercial available

[75, 77,78,79,80]

Size-based methods

Size- or molecular weight-dependent isolation

1. Ultrafiltration 2. Size exclusive chromatography (SEC) 3. Asymmetrical-flow field-flow fractionation (AF4)

Low (AF4)

High purity (AF4, SEC); exosomes may be deformed (ultrafiltration)

Low/moderate equipment cost (ultrafiltration and SEC ); high equipment cost (AF4)

1 - 1.5 h (Ultrafiltration); 0.5 - 1 h (SEC); 1h (AF4)

Available, .e.g.. EVSecond column (GL Sciences) or qEV column (Izon Science)

[82,83,84, 98,99,100]

Immunoaffinity-based methods

Antibody-antigen interaction-dependent isolation

1. Immunocapture 2. Immunoadsorption

Low

High purity

High reagent cost

4 - 5 h

Available, e.g. MagaCapture™ Exosome Isolation Kit (Wako) or Exosome-Human CD9 Isolation Reagent (ThermoFisher)

[85,86,87]

Precipitation-based methods

Solubility- or dispersibility-dependent isolation

Polyethylene glycol

Dependence

Contaminated precipitates

Low equipment cost

1 h

Available, e.g. Total Exosome Isolation Kit (Invitrogen) or ExoQuick™ Exosome Precipitation (System Biosciences)

[81, 88, 89]

Microfluidic-based methods

Size-, density-, or antibody-antigen interaction-dependent isolation

Microfluidic device with nanoporous membrane, immuno-chip, or porous silicon nanowires-on-micropillar structure

Dependence

High purity (porous nanowires-on-micropillar structure); EVs may be damaged (nanoporous membrane-based filtration)

Low/moderate equipment cost

2 h (porous membrane-based filtration); 1.5 h (immuno-chip)

No commercial available

[91,92,93,94,95]

Flow cytometry-based methods

Antibody-antigen interaction-dependent isolation

1. Fluorescent-labeled antibody-based isolation 2. Immuno-magnetic-based isolation

Low

High purity

High equipment and reagent cost

12 h

Avaialbe, e.g. Exosome Flow Cytometry Kit (Wako)

[96, 97]