Bioinspired microfluidic chip for fluorescent detection of ultrafast and stable PCR

Abstract We present a bioinspired microfluidic chip integrated with real-time optical detection for high-throughput polymerase chain reaction (PCR). The chip incorporates sloped-wall micropore arrays that serve dual functions: acting as droplet traps and optically active microcavities, thereby enhancing both light transmission and fluorescence signal collection. By optimizing the micropore geometry-specifically adjusting the depth-to-diameter ratio from 1:4 to 1:1 and the bottom diameter from 400 μm to 150 μm (as in Micropores 3–5), we achieved droplet arrays with a volume relative standard deviation below 10% and a filling efficiency above 97%. An optical detection module-comprising an LED source, bandpass filters, and a CMOS camera-was aligned with the chip for real-time fluorescence monitoring in 0.7 nl droplet array. To prevent evaporation under thermal cycling, a glass-PDMS-oil-glass multilayer architecture was employed, maintaining evaporation below 10%. Moreover, Triton X-100 surfactant effectively suppressed bubble formation. Compared to conventional 10 μl systems, this platform shortened PCR initiation time by 46% and plateau phase by 27%. This study highlights the bioinspired sloped-wall microstructures can synergistically improve droplet stability and optical signal acquisition, providing a promising strategy for compact, rapid, and robust nucleic acid detection.