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Small extracellular vesicles have been widely studied for their therapeutic properties and ability to deliver bioactive molecules. In addition to secretory vesicles, cells contain small intracellular vesicles involved in physiological and metabolic processes, whose therapeutic potential remains unexplored. Here, we developed protocols to isolate small intracellular vesicles from multiple cell types and systematically compared their molecular and functional profiles to extracellular vesicles. Intracellular vesicles are smaller, yield higher quantities, and demonstrate enhanced cellular uptake in both in vitro and in vivo models. Molecular profiling revealed that intracellular vesicles are enriched in proteins associated with the endoplasmic reticulum and Golgi apparatus, possess distinct microRNA signatures linked to intracellular membrane systems, and contain elevated levels of phospholipids such as phosphatidylcholine and phosphatidylethanolamine. Vesicles derived from umbilical cord mesenchymal stem cells showed superior therapeutic efficacy in a model of retinal degeneration by reducing endoplasmic reticulum stress and delivering neuroprotective factors. Additionally, intracellular vesicles exhibited enhanced drug-loading capacity and efficient delivery of lipophilic compounds to the retina. These findings position intracellular vesicles as promising candidates for therapeutic applications.