Extracellular vesicles (EVs) refer to vesicles that are released by cells into the extracellular space. EVs mediate cell-to-cell communication via delivery of functional biomolecules between host and recipient cells. EVs can be categorised based on their mode of biogenesis and secretion and include apoptotic bodies, ectosomes or shedding microvesicles and exosomes among others. EVs have gained immense interest in recent years owing to their implications in pathophysiological conditions. Indeed, EVs have been proven useful in clinical applications as potential drug delivery vehicles and as source of diagnostic biomarkers. Despite the growing body of evidence supporting the clinical benefits, the processes involved in the biogenesis of EVs are poorly understood. Hence, it is critical to gain a deeper understanding of the underlying molecular machineries that ultimately govern the biogenesis and secretion of EVs. This chapter discusses the current knowledge on molecular mechanisms involved in the biogenesis of various subtypes of EVs.