Nanofibrous Interface-Engineered Microgel Array Enabling Streamlined Digital PCR

Abstract

Digital polymerase chain reaction (dPCR) is increasingly used due to its key advantageshigh sensitivity, absolute quantification, and tolerance to contaminantsthat contribute to its overall robustness. However, it typically relies on complex droplet or microfluidic systems requiring pumps or vacuum systems for partitioning, limiting its accessibility and portability. Here, we present a microgel array chip comprising 2003 porous microgels (1 nL) in a 1 cm2 area, offering simple and rapid partitioning. Controlling internal porosity and surface structure of microgels is crucial to achieving reliable volume control, effective mass transfer, and amplification. In particular, the hydrophilic, oil-repelling nanofibrous architecture of the microgel surface enables reliable partitioning via sequential injection of the PCR mixture and oil. Its solid structure also provides stable amplification under fast thermal cycling, allowing the diagnostic assay to be completed in under 1 h. The platform demonstrates high accuracy in absolute quantification of nucleic acids, showing a strong correlation with conventional RT-qPCR in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nasopharyngeal swab samples. These results highlight the microgel array dPCR as a practical and accessible tool for rapid and accurate diagnostics.