Stimuli-responsive ‘On–Off’ SERS–darkfield bimodal plasmonic nanoprobes for selective cancer cell illumination

Early detection is essential for improving cancer treatment outcomes. Surface-enhanced Raman scattering (SERS) offers high sensitivity and molecular specificity but suffers from slow imaging speed when used independently. Here, we introduce stimuli-responsive bimodal SERS–dark-field (DF) nanoprobes for rapid and selective imaging of metastatic prostate cancer cells. These nanoprobes utilize legumain enzyme–responsive peptide coatings on plasmonic gold nanocubes and a bio-orthogonal self-condensation mechanism to facilitate enzyme-triggered nanoprobe aggregation. Upon internalization, the nanoprobes selectively aggregate in DU145 cells that overexpress legumain, while remaining uniformly dispersed in LNCaP cells with minimal legumain activity. DF microscopy quickly identifies these aggregates, significantly expediting subsequent SERS imaging. The engineered design of these nanoprobes ensures distinct vibrational signatures specifically correlated to enzyme activity, establishing a direct link between enzymatic presence and SERS signal generation. By combining the rapid localization capabilities of DF imaging with the molecular precision of SERS, our bimodal imaging strategy significantly surpasses conventional single-modal SERS techniques, providing a versatile platform for fast and accurate detection of various diseases through their distinctive enzymatic profiles.