Exosomes are a small type of extracellular vesicles containing nucleic acids, lipids, and proteins that are implicated in tumorigenesis, metastasis, and cardiac regeneration, and therefore serve as potentially useful biomarkers from fluids or as vehicles for drug delivery. Global bottom-up mass spectrometry-based proteomics has been previously used to profile exosome cargo for diagnostic purposes. However, the current protocols for MS analysis of extracellular vesicles and exosome proteomics are challenging due to labor-intensive sample preparation, including lengthy digestion times and removal of MS incompatible reagents, and the need for high sensitivity. To address these challenges, the present invention provides a novel, high-throughput strategy for extracellular vesicle analysis and exosome proteomics using a photo-cleavable, anionic surfactant, preferably 4-hexylphenylazosulfonate (Azo). These photo-cleavable, anionic surfactants are able to facilitate high-throughput digestion with minimal sample cleanup for quantitative and highly reproducible analysis.