Although animal-derived extracellular vesicles (EVs) are moving increasingly into scientific focus, EVs from other kingdoms remain underestimated and our knowledge of them is still expandable, probably due to the lack of an easy and broadly executable isolation, purification and visualization method. Using differential centrifugation with subsequent agarose gel electrophoresis, we were able to simplify the terms of EV isolation. EVs from Nicotiana tabacum L., Vinca minor L., and Viscum album L. were purified, even though they did not migrate into the gel matrix. If 3,3- Dihexyloxacarbocyanine iodide (DiOC 6 ) is added to the specimen in excess, membranous components can already be detected by eye, or with higher sensitivity, using a UV transilluminator. The sample preparation can be adjusted to the EV species of interest. Moreover, EVs are separated from small charged contaminants and dye excess, because these impurities can pass the gel matrix, while EVs themselves are retained in the pocket. Significantly, we isolated EVs from dried plant material, which is-to our knowledge-the first proof that EVs are stable enough to overcome the drying process of plant material.
Keywords: apoplastic fluid; dried plants; electrophoresis; extracellular vesicles; purification.