Every cell in the body secretes extracellular vesicles (EVs) possibly as cellular signaling components and these cell-derivatives can be found in multiple numbers in biological fluids. EVs have in the scientific field received great attention in relation to pathophysiology and disease diagnostics. Altered protein expressions associated with circulating EVs in diseased individuals can serve as biomarkers for different disease states. This capacity paves the way for non-invasive screening tools and early diagnostic markers. However, no isolation method of EVs has been acknowledged as the “golden standard,” thus reproducibility of the studies remains inadequate. Increasing interest in EV proteins as disease biomarkers could give rise to more scientific knowledge with diagnostic applicability. In this chapter, studies of proteins believed to be associated with EVs within cancer, autoimmunity, metabolic and neurodegenerative diseases have been outlined.