Extracellular vesicles (EVs) are nanosized, membrane-enclosed particles present in various body fluids, such as blood, saliva, cerebrospinal fluid, urine, and breast milk. Because their molecular content reflects the physiological and pathological status of their cells of origin, EVsEVs have emerged as powerful targets in liquid biopsyLiquid biopsy—a minimally invasive approach for early disease detection, prognosis, and therapeutic monitoring. Accurate characterization of EVs is critical for unlocking their diagnostic and therapeutic potential. This chapter focuses on immunoassaysImmunoassays—a central class of analytical tools that leverage the specific binding between antibodies and target proteins to detect and characterize EVs. Two main strategies to analyze the protein signature of EVs using immunoassays will be explored: (1) bulk methodsBulk methods allow simultaneous detection and semiquantitative analysis of large EV populations and biomarkers simultaneously and (2) single-EV analysisSingle-EV analysis focuses on profiling surface protein markers on individual vesicles, offering quantitative and high-resolution data using advanced techniques.