Hiroyuki Takei

Measurement of Intense Ultrasound Field in Air Using Fiber Optic Probe

In this report, a method of measuring intense ultrasound field using a fiber optic probe is studied. We try to utilize the modulation of optical reflectivity at the end of the optical fiber through a change in the refractive index of air due to sound pressure. The theoretical sensitivity for air is studied and a setup for the measurement is described. Comparisons with a commercial probe-type condenser microphone are performed. The absolute value of sound pressure estimated theoretically almost agreed with the results obtained using a conventional microphone and showed good linearity. The validity of the method was proved using three measurement examples.

Development of a high-speed real-time polymerase chain reaction system using a circulating water-based rapid heat-exchange

Polymerase chain reaction (PCR) is a powerful technique to detect microorganisms, viruses, or cells by amplifying a single copy or a few copies of a fragment of a particular DNA sequence. To reduce acquisition time, it is necessary to decrease the temperature transition time between denaturation and extension. We have developed a simple rapid real-time microlitter-sample droplet PCR system accomplished by the rapid liquid-based heat-exchange of sample droplets by quick switching of two circulating hot waters of denaturation and extension, a microlitter-sized droplet and a thin-film aluminum chip. Using this system, rapid PCR amplification of a set of droplets lined up on an aluminum chip was conducted successfully as shown by the increase in fluorescence intensity, and was accomplished within 3.5 min in 40 cycles of 1 s denaturation and 3 s extension reaction, which is one magnitude faster than conventional fast PCR systems. This method allows the rapid detection of DNA fragments and has a possibility for measuring multiple samples simultaneously in a miniaturized microfluidic chip.

Air Flow in a Small Gap between a Bending Vibrator and a Reflector

In this paper, we present numerical simulations and experiments of air flow induced in a gap between a piezoelectric bending vibrator and a reflector. Half of the simulation is based on the coupled analysis of piezoelectric elastic vibration and sound field using finite element method (FEM). The other half is the numerical fluid dynamics modeled using FEM where the driving forces are calculated on the basis of the acoustic streaming theory. One-directional streaming in the gap is predicted using the simulation and was successfully obtained in the experiments by introducing asymmetry both in the vibration distribution of the vibrator and the sound field. Air flow velocity of over 0.5 m/s was achieved using a vibrator of 30×20 mm2 with an electrical power consumption of 200 mW.

Cup-Shaped Superparamagnetic Hemispheres for Size-Selective Cell Filtration

We propose a new method of size separation of cells exploiting precisely size-controlled hemispherical superparamagnetic microparticles. A three-layered structure of a 2-nm nickel layer inserted between 15-nm silicon dioxide layers was formed on polystyrene cast spheres by vapor deposition. The polystyrene was then removed by burning and the hemispherical superparamagnetic microparticles, “magcups”, were obtained. The standard target cells (CCRF-CEM, 12 ± 2 μm) were mixed with a set of different sizes of the fabricated magcups, and we confirmed that the cells were captured in the magcups having cavities larger than 15 μm in diameter, and then gathered by magnetic force. The collected cells were grown in a culture medium without any damage. The results suggest that this method is quick, simple and non-invasive size separation of target cells.

Enhanced infrared LSPR sensitivity of cap-shaped gold nanoparticles coupled to a metallic film.

We report on optical properties of gold deposited on SiO2 nanospheres randomly adsorbed on a thin gold layer. Extinction peaks with optical density of more than 2 are observed in the visible as well as near-IR regimes. The peak wavelength of the latter was affected exquisitely by the thickness of the top layer. A helium ion microscope (HIM) was used for careful observation of morphological transformation accompanying the change in the deposition thickness. Growth of grain structures into a capped-dimer structure was accompanied by slight blue-shift of the visible peak and significantly greater red-shift of the near-IR peak. Our finite-difference time-domain (FDTD) calculations show that these peaks in the visible and near-IR can be respectively attributed to dipole modes associated with transverse and longitudinal oscillations of free electrons in the gold-capped dimer. To investigate the refractive index sensitivity of these peaks, we used two approaches: immersion in solutions of varying refractive index and coating with an organic layer. With the first approach that characterizes the bulk sensitivity, the visible peak shows sensitivity of 122 nm/RIU, while the near-IR peak shifts at the rate of 506 nm/RIU. With the second approach that reflects the local sensitivity, the surface was saturated with alkaline phosphatase (ALP), whose subsequent reaction led to formation of a thin insoluble organic layer, causing a relatively small blue-shift, under 7 nm, of the visible peak and much larger red-shift, over 50 nm, of the near-IR peak when measured in buffer. When the same reaction was measured at end points in the air, the shift was as large as 444 nm for the near-IR peak.

DNA Hybridization Efficiency on Concave Surface Nano-Structure in Hemispherical Janus Nanocups

We examined the effect of a concave structure on DNA hybridization efficiency using an inner surface of hemispherical Janus nanocups in the range from 140 to 800 nm. Target DNA was specifically immobilized onto the inner cup surface, hybridized with complementary DNA-attached 20 nm Au probes, and the number of the hybridized probes was counted by scanning electron microscopy. The hybridization density of the attached Au probes on 800 nm nanocups was 255 μm(-2), which was 0.57 times that on a flat surface, 449 μm(-2), and increased to 394 μm(-2) on a 140 nm cup, 0.88 times of a flat surface, as the cup size decreased. The local density of attached Au probes within the central 25% at the bottom of the 800 nm nanocups was 444 μm(-2), which was closer to that on a flat surface, and the tendency was the same for all sizes of cups, indicating that the size dependency of DNA hybridization efficiency on the concave structures were mostly affected by the lower efficiency of side wall hybridization.

Production of size-controlled nanoscopic cap-shaped metal shells

A method of producing precisely size-controlled metal nanoparticles is described. Polystyrene (PS) spheres placed on a substrate were used as a cast for the metal nanoparticles. The diameters of the PS spheres were processed into the desired sizes by oxygen plasma etching, and metal was deposited on the PS to the desired thickness by thermal evaporation. The PS casts were then removed by the UV-excited ozone oxidization reaction. The diameters of the obtained cap-shaped metal shells had a distribution within 5% of the coefficient of variation. These particles can be used as simultaneously applicable biological labels along with different-sized nanoparticles in immuno-electron microscopy.

TLC-SERS plates with a built-in SERS layer consisting of cap-shaped noble metal nanoparticles intended for environmental monitoring and food safety assurance

We report on a thin layer chromatograph (TLC) with a built-in surface enhanced Raman scattering (SERS) layer for in-situ identification of chemical species separated by TLC. Our goal is to monitor mixture samples or diluted target molecules suspended in a host material, as happens often in environmental monitoring or detection of food additives. We demonstrate that the TLC-SERS can separate mixture samples and provide in-situ SERS spectra. One sample investigated was a mixture consisting of equal portions of Raman-active chemical species, rhodamine 6 G (R6G), crystal violet (CV), and 1,2-di(4-pyridyl)ethylene (BPE). The three components could be separated and their SERS spectra were obtained from different locations. Another sample was skim milk with a trace amount of melamine. Without development, no characteristic peaks were observed, but after development, a peak was observed at 694 cm-1. Unlike previous TLC-SERS whereby noble metal nanoparticles are added after development of a sample, having a built-in SERS layer greatly facilitates analysis as well as maintaining high uniformity of noble metal nanoparticles.

Production of double-layered metal nanocups for artificial nanospace of biomolecular reaction

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.

Production of Nanoparticles Using Several Materials for Labeling of Biological Molecules

Various size-controlled metal nanoparticles (NPs) coated with probe DNAs have been developed. Gold, silver, germanium, copper, or nickel was thermally deposited as the inner layer on the surface of a polystyrene bead, and gold was coated as the outer layer for immobilizing thiolated probe DNAs by Au–S covalent bonding. The ultraviolet–visible (UV–vis) spectra of NPs showed that an outer gold layer thickness of 2 nm was sufficient for the immobilization of probe DNAs having a signal/noise (S/N) ratio of specific attachment of NP probes on the DNA chips eight-times higher than that of fluorescent probes. The size distributions of NPs were within the 6.7% coefficient of variation regardless of the type of metal and size. The different metal layers of NPs were also discriminated successfully by measuring backscattered electron intensity by scanning electron microscopy (SEM). The results indicate that NPs can be used for a two-dimensional probe set for SEM observation of size differences and differences in the type of metal used.