Table 3 |.

Microscopy methods

Modalities	Resolution (XY)	Resolution (Z)	Illumination	Probes	Acquisition time	Post-acquisition processing	Live or fixed
Standard fluorescence microscopy	250 nm	500 nm	Epi, confocal, TIRF	Conventional fluorescent probes	Seconds		Live, fixed
SIM, airyscan	80 nm–150 nm	250 nm–350 nm	Widefield (epi and TIRF)	Conventional fluorescent probes	Seconds	Yes, FTT	Live, fixed
STED	30 nm–80 nm	150 nm	Laser scanning	Limited selection of probes (match depletion laser)	Seconds	No	Live, fixed; optimal for fixed
Cryo-soft X-ray tomography	25 nm–40 nm	30 nm	Widefield	none		No	Fixed (near-native state vitrification)
PALM	20 nm	50 nm	Widefield (epi and TIRF)	Photoactivatible fluorescent proteins	Minutes	Yes (PSF mapping)	Live, fixed
STORM	20 nm	50 nm	Widefield (epi and TIRF)	Photoswitchable dyes	Minutes	Yes (PSF mapping)	Live, fixed
LLSM	100 nm–200 nm	400 nm	Multi-Bessel beam plane illumination	Conventional fluorescent probes	Seconds, minutes or hours	Not necessary, but often tracking dynamic processes	Live, fixed; optimal for live
TEM	<1 nm	70 nma	Electron beam	Contrast reagent, immunochemistry	Seconds	Yes	Fixed
CLEM	<1 nm/150 nmb	5 nmc	Electron beam, widefield	Contrast reagent, nanodots, and fluorescent proteins	Minutes	Yes (aligning)	Live and/or fixed

Characteristics of imaging methods used to visualize EVs. epi, epifluorescence; FTT, fast Fourier transform; PSF, point spread function; TEM, transmission electron microscopy.

^aResolution corresponding to the thickness of the section.

^bResolution gap between electron microscopy and light microscopy data, respectively.

^cTomography from double-tilted 250 nm sections.