Differential detergent sensitivity of extracellular vesicle subpopulations

Abstract

a Extracellular vesicles (including exosomes, microvesicles and apoptotic bodies) are currently attracting rapidly increasing attention from various fields of biology due to their ability to carry complex information and act as autocrine, paracrine and even endocrine intercellular messengers. In the present study we investigated the sensitivity of size-based subpopulations of extracellular vesicles to different concentrations of detergents including sodium dodecyl sulphate, Triton X-100, Tween 20 and deoxycholate. We determined the required detergent concentration that lysed each of the vesicle subpopulations secreted by Jurkat, THP-1, MiaPaCa and U937 human cell lines. We characterized the vesicles by tunable resistive pulse sensing, flow cytometry and transmission electron microscopy. Microvesicles and apoptotic bodies were found to be more sensitive to detergent lysis than exosomes. Furthermore, we found evidence that sodium dodecyl sulphate and Triton X-100 were more effective in vesicle lysis at low concentrations than deoxycholate or Tween 20. Taken together, our data suggest that a combination of differential detergent lysis with tunable resistive pulse sensing or flow cytometry may prove useful for simple and fast differentiation between exosomes and other extracellular vesicle subpopulations as well as between vesicular and non-vesicular structures.

Ceramide and Exosomes: A Novel Target in Cancer Biology and Therapy

“…Finally, loss-of-function assays will help exclude other sources of bioactive molecules in the exosomes preparation. These assays include UV irradiation (to destroy RNA in exosomes) or mild detergent treatment, which either obliterates the effect of exosomes (if based on RNA) or exosomes themselves (Coumans et al, 2017; Kinoshita et al, 2017; Osteikoetxea et al, 2015). Our group has specialized on pre-purification loss-of-function assays by preventing formation or secretion of exosomes in vitro and in vivo (Dinkins, Dasgupta, Wang, Zhu, & Bieberich, 2014; Dinkins, Enasko, et al, 2016; Kong, Hardin, et al, 2015; G.…”

Proteome profiling of extracellular vesicles captured with the affinity peptide Vn96: comparison of Laemmli and TRIzol© protein‐extraction methods

“…The use of TRIzol© for the isolation of nucleic acids from cells and EVs has been well established [11]; however, its utility for protein capture is less well investigated and has not been validated in the context of EVs. While work from the group of Edit Buzas [12] demonstrates differential detergent sensitivity of EV sub-populations, a similar effect was not anticipated with the TRIzol© protein-extraction reagent because TRIzol© (or Laemmli) extraction does not depend on this principle. The main barrier to using TRIzol© for protein recovery arises from poor reproducibility and difficulty in the re-solubilization of the protein pellet from the prescribed acetone- or isopropanol-mediated precipitation step [13].…”

Characterization of Membrane Integrity and Morphological Stability of Human Salivary Exosomes

“…Several other studies employing detergents reported on the stability of exosomes. [27][28][29] It would be interesting to compare the effects of detergents on the membrane integrity of exosomes prepared from various sources.…”