Viable Escherichia coli enumeration on a polydimethylsiloxane (PDMS) chip with vertical channel-well configuration

The culture-based methods for viable Escherichia coli (E. coli) detection suffer from long detection time and laborious procedures, whereas the molecule tests and immune recognition technologies lack live/dead E. coli differentiation. Rapid, easy-to-use, and accessible viable E. coli detection is of benefit to bacterial infection diagnosis and risk warning of E. coli contamination of water and food, safeguarding human health. Herein, we propose a microwell chip-based solution to realize simple and rapid determination of viable E. coli. The vertical channel-well configuration is applied to develop the microwell array chip for increasing the microwell density (6200 wells/cm²), yielding a broad dynamic range from 10³ to 10⁷ CFU/mL. We incorporate an inducible enzyme assay with the developed chip and achieve the differentiation of live/dead E. coli within 4 h, significantly shortening the detection time from over 24 h in the standard method. By encapsulating single E. coli into microwells, the concentration of viable cells can be determined simultaneously through counting positive microwells. In addition, the air soluble PDMS that can store negative pressure for independent sample digitalization endows the developed chip with simple operation and less reliance on external equipment. With further developments for increasing the number of microwell and integrating more sample panels, the developed chip can become a useful tool for rapid viable E. coli enumeration with user-friendly operation, simple procedures, and accessibility in decentralized settings, thereby deploying this device for water and food safety monitoring, as well as clinical bacterial infection diagnosis.