Morphological Characterization of Plasma-Derived Nanoparticles Isolated by High-Speed Ultracentrifugation: A Scanning Electron Microscopy Study

Extracellular vesicles are critical mediators of intercellular signaling. Recent studies have revealed that, in addition to vesicular structures, smaller non-vesicular nanoparticles-termed exomeres and supermeres-also participate in intercellular communication. Detailed characterization of these nanoscale entities within biological systems is essential for elucidating their structural and functional roles. Due to their sub-50 nm dimensions, high-resolution imaging modalities such as atomic force microscopy and electron microscopy are currently the primary techniques available for their visualization. In the present study, we employed low-voltage scanning electron microscopy to investigate the size of exomeres and supermeres isolated from human blood plasma via high-speed ultracentrifugation. Platelet-poor plasma was obtained from the blood of six healthy donors (two women and four men, aged 21-46 years). By ultracentrifugation (170,000× <i>g</i> for 4 h), the plasma was purified of extracellular vesicles. Two fractions were sequentially isolated: one containing exomeres (170,000× <i>g</i> for 20 h) and one containing supermeres (370,000× <i>g</i> for 20 h). The particles were examined using a Zeiss Auriga microscope with no sputter coating at an accelerating voltage of 0.4-0.5 kV. The images obtained from the fractions showed particles 10-50 nm in diameter, both individual particles and aggregated structures. The fractions were also slightly contaminated with larger particles, supposedly extracellular vesicles. Examining the fractions using a dynamic light scattering device additionally revealed the presence of particles 10-18 nm in size. It should be noted that the fractions obtained did indeed contain particles measuring 10-50 nm, which corresponds to the size of exomeres and supermeres. Low-voltage scanning electron microscopy allows for examination of the structure of exomeres and supermeres in blood plasma fractions. However, it should be noted that without the use of immunological identification, this method does not allow exomeres and supermeres to be distinguished from accompanying particles. It should also be noted that because the size of exomeres and supermeres is close to the detection threshold of low-voltage scanning electron microscopy, in such studies it is generally only possible to detect the size of these particles.