Microcavity Array-Based Digital SERS Chip for Rapid and Accurate Label-free Quantitative Detection of Live Bacteria

In this study, we developed a novel digital surface-enhanced Raman spectroscopy (SERS) chip that integrates an inverted pyramid microcavity array, a microchannel cover plate, and a multilayer gold nanoparticle (AuNP) SERS substrate. This innovative design exploits the synergistic effects of the microcavity array and the microchannel to enable rapid and large-scale digital discretization of bacterial suspensions. The concentration effect of the picoliter cavities, combined with the superior Raman enhancement effect of the multilayer AuNP SERS substrate, allows for the precise identification of live bacteria within the microcavities through in situ and label-free SERS testing after a short incubation period. By counting the resulting positive or negative signals, the concentration of the target analyte can be directly determined via Poisson statistics. Experimental results demonstrate that this method enables the accurate quantification of <i>Escherichia coli</i> (<i>E. coli</i>) BL21 within a 4-h incubation period. Compared with traditional analog SERS detection methods, our proposed digital SERS detection strategy reduces the impact of signal intensity fluctuations, thereby significantly improving detection efficiency and accuracy. We believe that this digital SERS chip has great application prospects in the fields of bacterial detection, antibiotic resistance analysis, and cellular dynamics monitoring.