Although the story of extracellular vesicles (EVs), these cell-derived submicron structures started about 70 years ago, we had to wait for the first transmission electron microscopic (TEM) evidence of their existence till 1981.
By now it has been proved that the release of membrane vesicles is a process conserved in both prokaryotes and eukaryotes. Compelling piles of evidence support the significance of exosomes, microparticles/microvesicles or ectosomes and apoptotic bodies in a broad range of biological processes including e.g. intercellular communication, signaling processes, the transfer of genetic information, and also in case of pathological events, from inflammation till tumor development. Not surprisingly, the possibility of EV-related biomarker and therapeutic applications made this research field attractive for clinical and pharmaceutical research, too.
However, in spite of growing interest, the classification of EVs is still under debate. The numerous protocols, applied by the different research teams, made the comparison of the results more complicated; moreover several frequently used and up-to-date particle enumeration techniques such as nanoparticle tracking analysis or tunable resistive pulse sensing are not suitable to distinguish vesicles and e.g. protein aggregates in similar size. Falsified experimental results may occur easily, especially in case of body fluid samples or culture media, in which protein aggregates, which share biophysical parameters with EVs, may contaminate vesicle preparations.
Purity and type of the fractions should be investigated with different techniques depending on the size range of the investigated vesicle population. For detection of exosomes, the frequently used flow cytometry is not the best choice, while by means of AFM, dynamic light scattering, MS techniques or microfluidic device, mentioned only few among the applied possibilities, can reveal several features of the investigated vesicle samples. However, even the most sophisticated combination of different techniques cannot be complete without transmission electron microscopy. It is the only technique which is able to visualize vesicular and non-vesicular particles in the whole size range of the vesicle population, and if we want to use EVs for research and therapy, we have to learn, how TEM can be applied for their accurate characterization.
Acknowledgements
This work was supported by grant NK 84043.
Figures:
Fig.1. Diverse population of microvesicles isolated from 5/4 cell culture supernatant
Fig.2. Human thrombocyte derived exosomes fraction following ultrafiltration. The fraction contains protein contamination (asterisk).
To cite this abstract:
Agnes Kittel, Xabier Osteikoetxea, Barbara Sódar, Krisztina Pálóczy, Tamás Baranyai, Zoltán Giricz, Edit Irén Buzás; Advantages and pitfalls for transmission electron microscopic studies in the identification of extracellular vesicles. The 16th European Microscopy Congress, Lyon, France. https://emc-proceedings.com/abstract/advantages-and-pitfalls-for-transmission-electron-microscopic-studies-in-the-identification-of-extracellular-vesicles/. Accessed: December 2, 2023« Back to The 16th European Microscopy Congress 2016
EMC Abstracts - https://emc-proceedings.com/abstract/advantages-and-pitfalls-for-transmission-electron-microscopic-studies-in-the-identification-of-extracellular-vesicles/