Table 2 Advantages and disadvantages of sEVs identification methods

Electron Microscope (EM)

Scanning electron microscopy or transmission electron microscopy

1. 1.It can directly observe the structure and morphology.

1. Sample pretreatment is complex and demanding.

2. Unable to accurately measure sample concentration after pretreatment.

[36]

Nanoparticle Flow Cytometry (NanoFCM)

Vesicles expressing specific protein were sorted using fluorescent antigen-antibody reactions.

1. Fast, Qualcomm volume.

2. The size and volume of the particles can be analyzed.

3. The influence of the contained impurities can be avoided.

1. Expensive equipment

2. Time-consuming

[37]

Dynamic light scattering (DLS)

Measuring fluctuation data of light intensity

1. High measurement sensitivity with lower limit of 10 nm.

2. Simple sample preparation and fast measurement speed.

1. The light intensity fluctuation signal of large particles will mask the light intensity fluctuation signal of small particles. It is not suitable for the measurement of complex samples with different sizes.

2. Unable to measure sample concentration.

[38]

Nanoparticle Tracking Analysis (NTA)

Measuring fluctuation data of light intensity

1. The nanoparticle can be directly and real-timely observed.

2. The mono-dispersity and poly-dispersity particles are more accurate.

The source of sEVs cannot be determined and may include fragments of membranes and other cells, or lipoprotein complexes.

[39]

Atomic Force Microscope (AFM)

Detecting an atomic interaction force between the sample surface and the element

1. Abundance, morphology, biomechanics and bio-molecular composition of sEVs can be characterized.

1. Expensive instrument

2. Small imaging range

[40]

Surface-enhanced Raman spectroscopy (SERS)

Detecting the scattered photons with the frequency changed due to the interaction between the incident light and the medium molecules

1. High sensitivity

2. High selectivity

3. Fast analysis

1. Reliable calibration model is needed and result processing is complex.

[41]

Protein determination

Antigen-antibody reaction

1. Simple operation.

1. Overall mean method, free protein cannot be distinguished.

2. Time-consuming

[42]