Nanocups (NCs), sub-micrometer semispherical bowls consisting of two different nanometer-thick metals on inner and outer layers, have been fabricated to mimic a localized nano-scale biochemical reaction environment for reactive biomolecules. Homogeneous polystyrene beads were used as a cast of the NCs, placed on a Si substrate, dried, and processed by oxygen plasma etching until the desired diameters and gaps among neighboring bead casts. For the fabrication of Au/Ni double-layered NCs, Au and Ni were sequentially deposited on upper halves of the bead surfaces by thermal evaporation with nanometer-order thickness control. The polystyrene casts were removed completely by UV-ozone oxidization reaction, and Au/Ni double-layered NCs were fabricated on a Si substrate. To orient the holes of the fabricated NCs to top for the substrate, poly(dimethylsiloxane) (PDMS) sol was dropped on the NCs placed on the Si substrate, hardened, and peeled off from the substrate, and then the NCs were placed on the PDMS surface with those holes turned-up. To examine the selective interaction of biomolecules on the inner layer of NCs as the artificial nanospace for biomolecular reactions, a thiolated target DNA was immobilized onto the inner layer of a Au/Ni NC as a model. The target DNA was labeled through hybridization reaction using small Au nanoparticles (NPs) on which a complementary probe DNA was immobilized. Both the surface-specific immobilization of the target DNA on the Au layer of the NC and the specific hybridization in NC nanospaces were confirmed by direct observations after those reactions using field emission scanning electron microscopy (FE-SEM), indicating that the inside of the fabricated NCs can be used as the artificial nanospace for studying localized biomolecular reactions.
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