Multiplex Optical Detection of Bacteria in Water Using a Cell Imprinted Polymer-Based Microfluidic Sensor

Abstract

Rapid detection of pathogenic bacteria in water is essential for environmental monitoring and public health. Here, we present a duplex microfluidic fluorescence sensor based on bacteria-specific cell-imprinted polymer (CIP) thin films for simultaneous detection of Salmonella enterica and Escherichia coli O157:H7. The sensor is fabricated using a simple, lithography-free approach, where CIP and nonimprinted polymer films are formed directly within laser-cut pressure-sensitive adhesive microchannels on a polymethyl methacrylate substrate. An orthogonal channel design creates six discrete sensing regions, enabling parallel and independent analysis. Detection is achieved through selective bacterial capture within imprinted cavities followed by FITC-based fluorescence readout. The sensor exhibits a concentration-dependent response over 104–107 CFU/mL, with a practical limit of detection of ∼105 CFU/mL. Competitive assays confirm high selectivity, with each CIP region responding preferentially to its target and minimal cross-reactivity in mixed-sample conditions. In addition, the sensor was successfully validated using Salmonella-spiked tap water samples, maintaining a concentration-dependent response, and demonstrating its applicability in realistic water matrices. The platform provides a low-cost, scalable, and portable solution for multiplex bacterial detection, with strong potential for field-deployable water quality monitoring applications.