A Large-field droplets for high-throughput Escherichia coli identification within one field of view

Droplet microfluidic chips have emerged as an efficient platform for single-cell analysis due to their high sensitivity, efficiency, and throughput, showing significant potential in pathogen detection. However, current droplet microfluidic chips encounter challenges in large-scale droplet quantification and precise imaging, rendering them unsuitable for the high-throughput pathogen detection required for a large number of samples. To address these issues, this study developed a high-precision fluorescence imaging system utilizing a confocal reflective fluorescence approach, which is an advanced microscopy technique that combines confocal microscopy and reflected fluorescence imaging. It can obtain fluorescence signal and reflected light signal in the sample at the same time, so as to provide richer and more comprehensive image information. The system offers a large field of view (17.8 mm × 17.8 mm) and high resolution (20 μm), enabling the rapid imaging of 30,000 droplets within 10 s, thereby significantly enhancing detection efficiency and automation. Additionally, the enzymatic reaction of Escherichia coli (E. coli) was implemented using the droplet microfluidic chip to validate the effectiveness of the optical imaging system, with results demonstrating the system's capability to accurately capture fluorescence changes during the reaction.