Optimization of Exosome Isolation from Vitis vinifera Cell Cultures

Plant extracellular vesicles (exosomes) are considered promising nanostructures for biomedical and biotechnological applications; however, methods for their isolation from plant cell cultures remain insufficiently optimized. The aim of this study was to evaluate the efficiency of exosome isolation from callus and suspension cultures of Vitis vinifera and to investigate the effects of culture age and extraction method on the yield of extracellular vesicles. It was demonstrated that suspension culture provides a higher exosome yield compared to callus tissue. Isolated particles were characterized by a size of 90-120 nm and a negative zeta potential, which corresponded to typical properties of plant extracellular vesicles. The maximum exosome yield was observed with a two-week-old suspension culture, which corresponded to the phase of intense cell growth. Comparison of extraction methods revealed that mechanical homogenization using a blender was the most efficient approach for exosome isolation, providing a high particle yield without significant deterioration in sample purity. The results demonstrate the potential of V. vinifera suspension cultures as a scalable platform for the production of plant-derived exosomes and may be useful for the development of biotechnological systems for the isolation of plant extracellular vesicles.