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. Author manuscript; available in PMC: 2024 Apr 25.
Published in final edited form as: Immunol Rev. 2022 Jul 8;312(1):76–102. doi: 10.1111/imr.13107

TABLE 3.

Exosome isolation methods

Advantages Disadvantages References
Ultracentrifugation (Most commonly used method) [Differential centrifugation (DC)]

  • Highly enriched EVs are produced.

  • Easiest and most widely used method.

  • Can analyze large volumes of samples.

  • Can analyze multiple samples at the same time.

  • Not clinically applicable as it is laborious, time-consuming, and low throughput.

  • Specific equipment and expertise are required.

  • The absolute separation of EVs is not possible.

  • EVs can be damaged due to exosome clumping.

  • EV Recovery is only 2% to 80%.

  • Cannot isolate pure EV.

 87,111,123–127
Precipitation

  • Six times faster than ultracentrifugation.

  • 2.5-fold higher concentration of exosomes per ml.

  • Highly reproducible.

  • Produces extracellular vesicles that have a significantly low number of both IgGs and albumin.

  • Clinically applicable.

  • EV recovery is 90%

  • Fast, inexpensive,

  • Requires no special equipment

  • Can be used for both low- and high-sample volumes.

  • Unable to isolate pure EVs.

  • Aggregation of exosomes.

  • Not suitable for identification of EV-associated biomarkers

 123–125,128
Size exclusion chromatography (SEC)

  • Clinically applicable.

  • Allows for size-based separation on a single column.

  • Can be used to obtain smaller size EVs.

  • EV recovery is 40%−90%.

  • Removes soluble components.

  • Processing time is higher.

  • Exosomes can be degraded due to buffer selection.

 124,125
Ultrafiltration (UF)

  • Faster method.

  • Superior to other methods for using large volumes of EV-containing fluids.

  • Remove soluble components & separate exosomes from big particles.

  • Inexpensive because of cheaper equipment.

  • Not clinically applicable.

  • Exosomes get attached to the filter pores and result in exosome loss.

  • Large vesicles can be damaged or deformed while passing through the membrane.

  • Cannot isolate pure EV.

 87,124,125,129–131
Immuno-capture assays (ICA)

  • Clinically applicable.

  • Simultaneously parallel ICA can be performed.

  • Isolate subpopulations of EVs.

  • Can efficiently detect members of the tetraspanin family, (e.g., CD81, CD9, and CD63).

  • Direct separation of exosomes from cell culture supernatant or bodily fluids is possible.

  • Highly specific.

  • High purity

  • Time-consuming (can take up to hours).

  • Cannot isolate large-volume samples.

  • Expensive equipment.

  • Low exosome yield.

 87,124,125,132,133
Density gradient centrifugation (DGC)

  • Can separate low-density exosomes from other EVs.

  • Separate EVs devoid of protein contaminants.

  • highly purified EVs.

  • Not clinically applicable as it is laborious, time-consuming, and low throughput.

  • Low exosome recovery rate.

 87,111,134,135
Polymer-based precipitation

  • Easy to use.

  • No need for specialized equipment.

  • Can be used for large samples.

  • Nonexosomal contaminants may be present.

  • Cannot isolate pure EV.

 136
Microfluidics

  • Rapid analysis is possible due to the short time.

  • Micro devices are used.

  • Low exosome yield.

  • Low reproducibility.

 124,137,138
Tangential flow filtration

  • Concentrate from large amounts of cell culture media.

  • Higher yield of EVs.

  • High reproducibility.

  • Produces exosome with fewer albumin contaminants.

  • Time-efficient.

  • Costly method due to expensive equipment and the use of disposable filters.

 139–142
Commercial kits miRCURY ExoQuick TEIR

  • Suitable alternatives to UC.

  • Similar zeta potentials to UC.

 143

Abbreviations: UC, Ultracentrifugation; TEIR, Total Exosome Isolation Reagent.