Extracellular vesicles (EVs) contain various glycans during their life cycle, from biogenesis to cellular recognition and uptake by recipient cells. EV glycosylation has substantial diagnostic significance in multiple health conditions, highlighting the necessity of determining an accurate glycosylation pattern for EVs from diverse biological fluids. Reliable and accessible glycan detection techniques help to elaborate the glycosylation-related functional alterations of specific proteins or lipids. However, multiple obstacles exist, including the inconsistency in glycosylation patterns between an entire batch of EVs and a specific EV protein, and difficulty in distinguishing glycosylation types after tedious separation and purification procedures. This review outlines recent advances in EV glycan detection, either at the glycomic level for a collection of intact EVs or at the molecular level for a specific protein on EVs. Particular emphasis has been placed on the abundance of EVs in body fluids and their unique characteristics for drug delivery of EVs, indicating an opportunity for diagnostic and therapeutic purposes via EV glycans.