In recent years, knowledge of cell-released extracellular vesicle (EV) functions has undergone rapid growth. EVs are membrane vesicles loaded with proteins, nucleic acids, lipids, and bioactive molecules. Once released into the extracellular space, EVs are delivered to target cells that may go through modifications in physiological or pathological conditions. EVs are nano shuttles with a crucial role in promoting short- and long-distance cell–cell communication. Comprehension of the mechanism that regulates this process is a benefit for both medicine and basic science. Currently, EVs attract immense interest in precision and nanomedicine for their potential use in diagnosis, prognosis, and therapies. This review reports the latest advances in EV studies, focusing on the nature and features of EVs and on conventional and emerging methodologies used for their separation, characterization, and visualization. By searching an extended portion of the relevant literature, this work aims to give a summary of advances in nanomedical applications of EVs. Moreover, concerns that require further studies before translation to clinical applications are discussed.