Table 2.

EV characterization techniques’ advantages and drawbacks.

EV Characterization Techniques	Aims	Advantages	Drawbacks
Dynamic light scattering (DLS)	Identifying EV distribution and size [87]	The best method for measuring a single type of nanoparticle in a solution and is easy to be performed [96]	Does not allow for the nanoparticles to be visualized [96]
Nanoparticle tracking analysis (NTA)	Identifying EV distribution and size [91]	Determination of very small EVs up to 30 nm, sample preparation does not affect the morphology of EVs, very quick and easy sample preparation, and samples can be reused after the measurements and can detect fluorescently labeled antibodies targeting EVs antigens [87,92]	Masking of small size particle by large size particles, as in DLS, finding the most appropriate dilution factor to obtain resonant results, and the fluorescent signal should be very bright to detect the EV phenotype accurately. Therefore, it is recommended to use antibodies coupled with quantum dots (Q-dots), which are very bright fluorochromes [87,92]
Transmission electron microscopy (TEM)	Identifying EV form [93]	The interior morphology of a particle may be seen via TEM, which can also reveal details on the size of the particle [97]	Complex sample preparation (numerous processes and EV morphological alteration). Biological specimens can be destroyed [93].
Western blot (WB)	Identifying EV marker protein expression [83,84,85]	Evaluates marker proteins in both qualitative and quantitative ways [97]	Complicated and consumes an extended period [29].
Flow cytometry	Identifying EV biomarkers [87]	Provides high-speed analysis and needs minimal sample concentration [97]	Takes a lot of time and is very complicated [97]