Table 2.

Evaluation of strategies for cargo loading into EVs

Strategies	Methods	Principles	Advantages	Limitations	References
Endogenous strategy	Gene transfection	Liposomes-mediated cargos delivery	Simple, no damage to membrane integrity	Cytotoxicity, low loading efficiency	[63, 64]
	Co-incubation	Penetration of cargos into cells	Simple, no damage to membrane integrity	Low specificity, low loading efficiency	[65, 66]
Exogenous strategy	Incubation	Interaction between cargos and EV membrane	Simple and feasible	Low loading efficiency, toxicity	[68–70]
	Electroporation	Transient voltage-induced the generation of pores on EV membrane	Simple, high loading efficiency	EV aggregation, nucleic acids precipitation	[71–73]
	Sonication	Deformation of EV membrane	High loading efficiency	EV aggregation	[74–77]
	Extrusion	Mechanical force-induced the temporarily destroyed membrane of EVs	Efficient packaging	May compromise the membrane structure of EVs	[78–80]
	Freeze–thaw cycles	Ice crystals-induced changes in EV membrane	High loading efficiency	EV aggregation, may change the membrane structure of EVs	[81–83]