A novel and efficient method to produce surface enhanced Raman scattering (SERS)-active nanogaps was developed by double-step Au film depositions and subsequent thermal annealings. The obtained nanostructure comprises metallic sea, islands, and in particular gaps between the sea and islands whose width can be automatically adjusted <10 nm, which is difficult to be achieved by the conventional single-step metal deposition and annealing. The nanogap structure gave an increased enhancement factor up to the order of 10 9 , whereas conventional vacant sea and Au island structure exhibited 10 7 . The SERS-active performances are supported by increased localized surface plasmon at the nanogaps.

Human-induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) are expected to have applications in the fields of regenerative medicine and drug discovery. However, the immaturity of hiPS-CMs is an issue to be considered, and in order to replicate in vivo responsivity, there have been several attempts to induce maturation in hiPS-CMs, including methods to induce differentiation of hiPS-CMs by changing culture medium and culture substrate. In particular, anisotropic culture, in which the cultured cells are aligned in one direction, induces the cellular morphology resembling that of in vivo cardiomyocytes and is expected to be a useful method for maturation of hiPS-CMs. We tried forming a nanostructure on the surface of the cell culture substrate using our original nanofabrication technology, with the aim of aligning cardiomyocytes and inducing maturation. Our newly developed nanostructure for anisotropic culture (line/dot structure) comprises a region of cone-shaped nanopillars with pitch distance of several hundred nm and a planar region, alternating in a striped pattern with intervals of several tens of µm, arranged on the surface of the cell culture substrate. The hiPS-CMs cultured using the line/dot structure showed anisotropic orientation, and increased mRNA expression was observed in myocardial structural protein genes, genes relating to ion channels, and the gene for Cx43. These results suggest that the line/dot nanostructure is effective for anisotropic culture and cell maturation of hiPS-CMs.