Table 2 Current techniques for the isolation of extracellular vesicles and their comparison
From: Extracellular vesicles: a rising star for therapeutics and drug delivery
Isolation techniques (Mechanism of isolation) | Advantages | Disadvantages | Isolation time | Yield and purity | Refs. |
|---|---|---|---|---|---|
Ultracentrifugation (Size; Density) | Simple; Cheap; Gold standard | Vulnerable to contamination by protein aggregates; Time-consuming; High demand for sample volumes; Expensive instruments; Low recovery rate; Structures easily damaged | > 4 h | Low yield and Low purity | |
Density gradient ultracentrifugation (Size; Density) | High purity; Exosome subpopulations can be isolated | Time-consuming; Larger losses; Cumbersome operation | > 16 h | Low yield and High purity | |
Size exclusion chromatography (Size; Molecular weight) | Structural integrity; Low usage; Saves time and effort; Ability to isolate specific subgroups of EVs | Wider size distribution; Contaminants such as protein aggregates and lipoproteins; Special columns required | 10–20 min per sample | High yield and High purity | |
Ultrafiltration (Size; Molecular weight) | Simple; Low cost; Variable sample injection volume; Rapid | Easy to cause clogged pores; EVs are adsorbed on the filter surface; Leading to a loss of yield; Shear forces may damage EVs | ~ 1 h for 200 mL cell culture media | High purity | |
Field-flow fractionation (Size; Molecular weight) | Label-free; Gentle; Rapid; Highly reproducible; High resolution | Small sample capacity; Analytes need to be stratified and concentrated beforehand; Samples need to be graded according to sample size | < 1 h | High purity | |
Precipitation-based methods (Solubility; Charge) | Easy to operate; Commercial kits are available; No specific equipment required | Protein aggregates may be precipitated; Commercial kits are expensive | 0.3–12 h | High yield and Low purity | |
Microfluidics (Affinity; Density; Size; Acoustic; Electrophoretic) | Fast; Low sample consumption; High recovery rate; High yield; automation; High portability | High cost; need for external force; Not suitable for large-scale production and requires method validation; Sample may evaporate | < 1 h | High yield and High purity | |
Affinity-based methods (Affinity) | High specificity; Rapid | Cumbersome process; Long operation time; High cost; Not suitable for large-scale production; Low yield; Requires subsequent isolation and purification steps | 4–20 h | Low yield and High purity |