Exosomes are small extracellular vesicles important to intercellular communication. Formed when multivesicular bodies (MVB) fuse with the plasma membrane, exosomes are released into the cellular environment where they can deliver their cargo. Many exosome cargoes, including proteins, mRNA, and lipids, are key for cell-to-cell and tissue-to-tissue crosstalk, such as between muscle and adipose. While involved in normal intercellular signaling, exosomes are also emerging as key players in a variety of disease phenotypes (e.g., the release of enlarged, specialized exosomes from tumor cells). Despite increases in exosome research in the past several decades, several key questions remain. Which proteins are involved in exosome biogenesis, and how are cargo targeted to exosomes? What role(s) could exosomes play in disease diagnosis and treatment? How can exosomes be harnessed for use in the lab or the clinic? In this review, we will discuss what is currently known about exosome biogenesis and cargo sorting, as well as how this exosome research is fueling the development of new technologies and therapeutics.