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Effective capture and rapid detection of pathogenic bacteria causing pandemic/epidemic diseases is an important task for global surveillance and prevention of human health threats. Here, we present an advanced approach for the on-site capture and detection of pathogenic bacteria through the combination of hierarchical nanostructures and a nuclease-responsive DNA probe. The specially designed hierarchical nanocilia and network structures on the pillar arrays, termed 3D bacterial capturing nanotopographical trap, exhibit excellent mechanical reliability and rapid (<30 s) and irreversible bacterial capturability. Moreover, the nuclease-responsive DNA probe enables the highly sensitive and extremely fast (<1 min) detection of bacteria. The bacterial capturing nanotopographical trap (b-CNT) facilitates the on-site capture and detection of notorious infectious pathogens (Escherichia coli O157:H7, Salmonella enteritidis, Staphylococcus aureus, and Bacillus cereus) from kitchen tools and food samples. Accordingly, the usefulness of the b-CNT is confirmed as a simple, fast, sensitive, portable, and robust on-site capture and detection tool for point-of-care testing.

Herein, we present a large-area 3D hemispherical perforated microwell structure for a bead based bioassay. Such a unique microstructure enables us to perform the rapid and stable localization of the beads at the single bead level and the facile manipulation of the bead capture and retrieval with high speed and efficiency. The fabrication process mainly consisted of three steps: the convex micropatterned nickel (Ni) mold production from the concave micropatterned silicon (Si) wafer, hot embossing on the polymer matrix to generate the concave micropattened acrylate sheet, and reactive ion etching to make the bottom holes. The large-area hemispherical perforated micropatterned acrylate sheet was sandwiched between two polydimethylsiloxane (PDMS) microchannel layers. The bead solution was injected and recovered in the top PDMS microchannel, while the bottom PDMS microchannel was connected with control lines to exert the hydrodynamic force in order to alter the flow direction of the bead solution for the bead capture and release operation. The streptavidin-coated microbead capture was achieved with almost 100% yield within 1 min, and all the beads were retrieved in 10 s. Lysozyme or thrombin binding aptamer labelled microbeads were trapped on the proposed bead microarray, and the in situ fluorescence signal of the bead array was monitored after aptamer-target protein interaction. The protein-aptamer conjugated microbeads were recovered, and the aptamer was isolated for matrix assisted laser desorption/ionization time-of-flight mass spectrometry analysis to confirm the identity of the aptamer.