This study introduces a novel method, the lectin-based thermophoretic assay (EVLET), for profiling extracellular vesicles (EVs) in plasma samples from triple-negative breast cancer (TNBC) patients. EVLET combines vibrating membrane filtration (VMF) and thermophoretic amplification to rapidly, sensitively, and cost-effectively analyze EV glycans. The method identifies TNBC-associated glycans using bioinformatics and selects a panel of lectins that can bind to these glycans. The EVLET system was validated in a pilot cohort study, achieving 91% accuracy for TNBC detection and 96% accuracy for longitudinal monitoring of therapeutic response. The EV glycan signature, representing the weighted sum of three lectins, was also found to predict TNBC progression with high accuracy. This study lays the foundation for non-invasive cancer management using EV glycans.This study introduces a novel method, the lectin-based thermophoretic assay (EVLET), for profiling extracellular vesicles (EVs) in plasma samples from triple-negative breast cancer (TNBC) patients. EVLET combines vibrating membrane filtration (VMF) and thermophoretic amplification to rapidly, sensitively, and cost-effectively analyze EV glycans. The method identifies TNBC-associated glycans using bioinformatics and selects a panel of lectins that can bind to these glycans. The EVLET system was validated in a pilot cohort study, achieving 91% accuracy for TNBC detection and 96% accuracy for longitudinal monitoring of therapeutic response. The EV glycan signature, representing the weighted sum of three lectins, was also found to predict TNBC progression with high accuracy. This study lays the foundation for non-invasive cancer management using EV glycans.