Extracellular vesicles are a heterogeneous population of membrane vesicles, released from cells both in vivo and in vitro, with an important role in intercellular communication. They have been isolated from a variety of biological samples such as blood, bronchoalveolar lavage, synovial fluid, urine, amniotic fluid, semen, milk, saliva; and from cell cultures of most mammalian cells. Based on their size and the site of formation in the cell, they are classified as exosomes (30–100 nm), microvesicles (100–1,000 nm) and apoptotic bodies (1,000–5,000 nm). Their protein, nucleic acid (miRNA) and lipid composition reflects the composition ofthe parental cell and depends on the current state ofthe cell. Consequently, the extracellular vesicles are an important source of novel biomarkers for diverse physiological conditions with important diagnostic and prognostic values, especially for disorders of the central nervous system. Researchers also investigate the potential use of extracellular vesicles as targets for diagnostic and therapeutic purposes, but many obstacles still have to be overcome. Due to their great clinical potential, the interest in extracellular vesicle characterization, formation and role increased importantly in recent years, fueling also development of new methods for their isolation and analysis. In the present review, we will describe the types of extracellular vesicles, methods for their isolation and analysis, their biological role and clinical potential.