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. 2021 Jun 2;11(6):1476. doi: 10.3390/nano11061476

Table 2.

Summary of selected papers in the literature studying EV structure with SAS and diffraction techniques.

Paper Aim Sample/Extraction Technique
(Q-Range nm−1)		 Model Main Findings
Varga et al.,
2014 [121]		 Investigating biophysical properties, i.e., shape and size distribution, of EVs isolated from erythrocytes. Erythrocyte-derived EVs/RBCs were removed by 2 centrifugation steps at 1550× g, t = 20 min and 20 °C. Next, the supernatant was centrifuged (18,890× g, 30 min) to concentrate EVs SAXS
(0.015–2.5)		 Scattering intensity comprises three contributions:

  • EV scattering modelled with a core–shell form factor weighted with a log-normal distribution;

  • Protein contribution modelled with

    spherical form factor;

  • Constant background scattering.

 Proper modelling of the scattering curve enabled obtaining the size distribution of EVs and discerning EV scattering from contaminants (which can co-precipitate during the purification process).
Romancino et al.,
2018 [123]		 Exploring the structural arrangement of the lipid bilayer of EV membranes with altered S-palmitoylation state. EVs secreted by skeletal muscle cells (C2C12 myotubes) at the 3rd day of differentiation (untreated and treated to inhibit S-palmitoylation)/
Ultracentrifugation at 118,000× g for 70 min		 SAXS
(0.03–6.0)
SANS
(0.05–4.0)		 Model-free analysis of SAXS and SANS profiles with neutron contrast variation. Analysis of a hump in the SAXS/SANS scattering profile centered at approximately q = 1.2 nm−1, which provide structural information on the bilayer organization (2π/q = 5.2 nm) SAXS and SANS with neutron contrast variation enables detecting subtle changes in the lipid membrane arrangement in terms of phospholipid head groups and hydrophilic tails associated with the S-palmitoylation state.
Montis et al.,
2020 [124]		 Studying the interaction between EV-derived supported lipid bilayers (EVSLBs) and gold-coated superparamagnetic iron oxide nanoparticles (SPIONs). Results were compared with artificial SLBs. EVs secreted by murine prostatic tumor cells (TRAMP-C2 cell line)/
Ultracentrifugation at 100,000× g for 240 min		 XRR
(0.15–0.25)
GISAXS
(0.15–0.25)
  • (i.)

    Model-free analysis of a specific signature of the GISAXS pattern, providing information on the EVSLB/SPION interaction;

  • (ii.)

    XRR curves were modelled as a multilayer composed of four layers, (i.e., inner polar group, lipid chain, outer polar headgroup, and SPIONs), each characterized by its thickness, scattering length density, and roughness.

 As measured with GISAXS, SPIONs are simply absorbed on both SLB surfaces, without membrane/nanoparticle reorganization and thus, without altering membrane biomechanics. A higher absorption is observed on the EVSLBs compared to POCP-SLB, as a consequence of its higher roughness associated with the protein content of exosomes, as measured with XRR.
Accardo et al.,
2013 [122]		 Classifying exosomes obtained from healthy and cancer cells and concentrated on superhydrophobic patterned surfaces. Exosomes extracted from two different CCD841-CoN (healthy epithelial colon) cell line and HCT116 (colorectal cancer) cell lines/
ExoQuick Precipitation Solution		 WAXS
(0.0–3.0)
SAXS
(0.0–1.8)		 Model-free analysis of micro-WAXS/SAXS lamellar peaks in the 3.5 nm−1 q range.
Micro SAXS patterns measured with benchtop instruments were deionized with a restoration algorithm.		 Micro-SAXS/WAXS measurements highlighted differences in the exosome macroaggregates morphology (i.e., number of orders, periodicity, and peak broadening). The authors hypothesized this was due to a more regular organization of exosomes derived from cancer cells than those one extracted from healthy cells, which could be useful to distinguish exosomes with different origins, also for diagnostic purposes.