At present, iron oxide nanoparticles (IONPs) are widely used in the biomedical field. They have unique advantages in targeted drug delivery, imaging and disease treatment. However, there are many things to pay attention to. In this paper, we reviewed the fate of IONPs in different cells and the influence on the production, separation, delivery and treatment of extracellular vesicles. It aims to provide cutting-edge knowledge related to iron oxide nanoparticles. Only by ensuring the safety and effectiveness of IONPs can their application in biomedical research and clinic be further improved.