Isolation and Characterization of Extracellular Vesicles from Natural Killer Cells

Natural killer (NK) cells are critical components of the immune system, recognized for their ability to identify and eliminate target cells undergoing various forms of stress. Beyond their direct cytotoxic activity, NK cells secrete extracellular vesicles (EVs), which serve as mediators of intercellular communication. These vesicles play an essential role in modulating immune responses and influencing tumoral and post-infection microenvironments. In this study, we present a detailed methodology for the isolation and expansion of human primary NK cells, the accumulation of their EVs, and the subsequent procedures for their isolation and characterization. © 2025 Wiley Periodicals LLC. Basic Protocol 1: NK cell isolation and expansion Basic Protocol 2: NK-EV purification using ultracentrifugation Alternate Protocol: NK-EV isolation using size exclusion chromatography Basic Protocol 3: Characterization of NK-EVs.