Detection of Biomarkers for EMT-Associated Phenotype Switching in Melanoma-Derived Extracellular Vesicles

Melanoma cells exhibit epithelial-to-mesenchymal transition (EMT)-like phenotype switching during targeted therapy, which contributes to the development of drug resistance. Extracellular vesicles (EVs) released by melanoma cells carry surface proteins reflective of this EMT-associated transition, providing an accessible window to track therapeutic response. This chapter presents a step-by-step protocol for profiling EMT-related melanoma EV protein signatures directly from unpurified sera using a microfluidic surface-enhanced Raman scattering (SERS) biosensing platform. The method integrates melanoma-specific EV capture, alternating current-electrohydrodynamic nanomixing-enhanced binding, and multiplex SERS nanotag labeling to enable sensitive quantification of EMT surface markers in a streamlined assay. Detailed instructions for reagent preparation, SERS nanotag synthesis, biosensor fabrication and operation, spectral acquisition, and data processing are provided to support implementation in melanoma translational research laboratories. This protocol enables reproducible monitoring of EMT-like phenotypic evolution in melanoma models and patient samples and may assist future studies aiming to predict or mechanistically dissect resistance to targeted therapy.