We present an analysis of the results of in situ surface-enhanced Raman scattering (SERS) of bacteria using a microfluidic chip capable of continuously sorting and concentrating bacteria via three-dimensional dielectrophoresis (DEP). Microchannels were made by sandwiching DEP microelectrodes between two glass slides. Avoiding the use of a metal nanoparticle suspension, a roughened metal surface is integrated into the DEP-based microfluidic chip for on-chip SERS detection of bacteria. On the upper surface of the slide, a roughened metal shelter was settled in front of the DEP concentrator to enhance Raman scattering. Similarly, an electrodepatterned bottom layer fabricated on a thin cover-slip was used to reduce fluorescence noise from the glass substrate. Gram positive (Staphylococcus aureus) and Gram negative (Pseudomonas aeruginosa) bacteria were effectively distinguished in the SERS spectral data. Staphylococcus aureus (concentration of 106 CFU/ml) was continuously separated and concentrated via DEP out of a sample of blood cells. At a flow rate of 1 μl /min, the bacteria were highly concentrated at the roughened surface and ready for on-chip SERS analysis within 3 min. The SERS data were successfully amplified by one order of magnitude and analyzed within a few minutes, resulting in the detection of signature peaks of the respective bacteria.
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