Isolation and Biophysical Characterization of Extracellular Vesicles from Hairy Root Cultures

Extracellular vesicles (EVs) are membrane-bound, non-replicating particles released by virtually all types of cells. EVs concentrate and deliver a plethora of biomolecules driving very important biological functions, including intercellular communication not only between cells of the same organism but also across different kingdoms. Plant extracellular vesicles (PEVs) are a promising alternative to mammalian EVs in biomedical applications. Here, we present an optimized and reproducible protocol for isolating PEVs from the hairy root (HR) cultures of medicinal plants Salvia dominica and S. sclarea. Our methodological approach introduces a significant advancement in the standardization of HR-EVs purification processes from plant biotechnological platforms, paving the way for their broader application across various sectors, including agriculture, pharmaceuticals, and nutraceuticals. Key features • Hairy roots serve as a plant biotechnological platform for the production of bioactive compounds. • Plant extracellular vesicles (EVs) provide a safer alternative to mammalian EVs for biomedical applications. • This protocol represents a significant advancement in the standardization of plant EV purification.