A microfluidic device enabling the isolation and concentration of bacteria for analysis by confocal Raman spectroscopy is presented. The glass-on-silicon device employs a tapered chamber surrounded by a 500 nm gap that serves to concentrate cells at the chamber apex during sample perfusion. The sub-micrometer gap retains bacteria by size exclusion while allowing smaller contaminants to pass unimpeded. Concentrating bacteria within the fixed volume enables the use of single-point confocal Raman detection for the rapid acquisition of spectral signatures for bacteria identification. The technology is evaluated for the analysis of E. cloacae, K. pneumoniae, and C. diphtheriae, with automated peak extraction yielding distinct spectral fingerprints for each pathogen at a concentration of 103 CFU/ml that compare favorably with spectra obtained from significantly higher concentration reference samples evaluated by conventional confocal Raman analysis. The nanogap technology offers a simple, robust, and passive approach to concentrating bacteria from dilute samples into well-defined optical detection volumes, enabling rapid and sensitive confocal Raman detection for label-free identification of focused cells.
© 2023 Author(s).