アプタマーを介した捕捉とチップ上での消化による高効率エクソソームプロテオミクスのための統合ピペットチッププラットフォームCLCtips

Current extracellular vesicles (EVs) isolation and proteomics preprocessing workflows are often complex, time-consuming, and costly and frequently rely on specialized equipment. Here, we present an integrated Capture-Lysis-Capture pipette-tips (CLCtips) platform that combines CD63-aptamer-mediated capture of EVs with epoxy-based protein immobilization and on-tip digestion. In our workflow, EVs-enriched fractions are first obtained from plasma, urine, and cerebrospinal fluid (CSF) by short size-exclusion chromatography and are then processed in CLCtips. This single-tip platform seamlessly integrates EVs enrichment, lysis, protein immobilization, and digestion into a 3 h workflow that requires only standard centrifugation equipment. The CLCtips were further applied to EVs-enriched fractions extracted from different biofluids, demonstrating high sensitivity by identifying an average of 2,388 proteins from 1 μL plasma, 1,034 proteins from 10 μL CSF, and 2,535 proteins from 20 μL urine (n = 3). The easy-to-use CLCtips process minute volumes with consumables costing below $1.2 per sample, and its modular architecture allows for customization with alternative affinity ligands, suggesting broad applicability in biomedical research areas.