Table 1.

Pros and cons of different EV isolation procedures.

Isolation Procedure	Pros	Cons	References
Differential ultracentrifugation (UC)	Still considered the gold standard for exosome isolation; less expensive than DG-UC	Costly equipment; high volume requirement; possible mechanical damage and incomplete EV isolation from contaminants	[26]
Density gradient ultracentrifugation (DG-UC)	High recovery Elevated EV purity and integrity	Pre-purification of samples; time-consuming procedure	[27]
Ultra-filtration (UF) + size exclusion (SE) chromatography	Suitable for analyzing large volumes. Fast and high recovery efficiency	Requires an appropriate choice of filter pore size and further purification	[28]
EV–polymer interaction/precipitation (A) or immuno- affinity capture by antigen/antibody binding (B)	Quick and easy; (A and B): EV recovery higher than UC.	(A) Low purity, protein contaminants. (B) Expensive, not suitable for large samples and EV sorting by size	[29]
Microfluidic techniques (±optical or spectroscopic devices) using a passive approach by passive filtration through nano-porous membranes (A) active approach exploiting physical forces applied to EV-containing fluids or suitable substrates coupled to antibodies for selected EV antigens (B)	Membranes remove cell debris allowing the sEV passage; high recovery Ability to analyze small volumes; quick and high recovery	Figure 1 Potential damage to EVs due to shear stress A and B still require validation and standardization	[23,30,31,32,33] [34,35]