Edge-Topological Regulation for in Situ Fabrication of Bridging Nanosensors.

In situ fabrication of well-defined bridging nanostructures is an interesting and unique approach to three-dimensionally design nanosensor structures, which are hardly attainable by other methods. Here, we demonstrate the significant effect of edge-topological regulation on in situ fabrication of ZnO bridging nanosensors. When employing seed layers with a sharp edge, which is a well-defined structure in conventional lithography, the bridging angles and electrical resistances between two opposing electrodes were randomly distributed. The stochastic nature of bridging growth direction at the sharp edges inherently causes such unintentional variation of structural and electrical properties. We propose an edgeless seed layer structure using a two-layers resist method to solve the above uncontrollability of bridging nanosensors. Such bridging nanosensors not only substantially improved the uniformity of structural and electrical properties between two opposing electrodes but also significantly enhanced the sensing responses for NO2 with the smaller variance and the lower limit of detection via in situ controlled electrical contacts.

Image Processing and Machine Learning for Automated Identification of Chemo-/Biomarkers in Chromatography-Mass Spectrometry.

We present a method named NPFimg, which automatically identifies multivariate chemo-/biomarker features of analytes in chromatography-mass spectrometry (MS) data by combining image processing and machine learning. NPFimg processes a two-dimensional MS map (m/z vs retention time) to discriminate analytes and identify and visualize the marker features. Our approach allows us to comprehensively characterize the signals in MS data without the conventional peak picking process, which suffers from false peak detections. The feasibility of marker identification is successfully demonstrated in case studies of aroma odor and human breath on gas chromatography-mass spectrometry (GC-MS) even at the parts per billion level. Comparison with the widely used XCMS shows the excellent reliability of NPFimg, in that it has lower error rates of signal acquisition and marker identification. In addition, we show the potential applicability of NPFimg to the untargeted metabolomics of human breath. While this study shows the limited applications, NPFimg is potentially applicable to data processing in diverse metabolomics/chemometrics using GC-MS and liquid chromatography-MS. NPFimg is available as open source on GitHub (http://github.com/poomcj/NPFimg) under the MIT license.

Maximizing Conversion of Surface Click Reactions for Versatile Molecular Modification on Metal Oxide Nanowires.

Click reactions (e.g., Huisgen cycloaddition) on metal oxide nanostructures offer a versatile and robust surface molecular modification for various applications because they form strong covalent bonds in a wide range of molecular substrates. This study reports a rational strategy to maximize the conversion rate of surface click reactions on single-crystalline ZnO nanowires by monitoring the reaction progress. p-Polarized multiple-angle incidence resolution spectrometry (pMAIRS) and Fourier-transformed infrared (FT-IR) spectroscopy were employed to monitor the reaction progress of an azide-terminated self-assembled monolayer (SAM) on single-crystalline ZnO nanowires. Although various reaction parameters including the concentration of Cu(I) catalysts, triazolyl ligands, solvents, and target alkynes were systematically examined for the surface click reactions, 10-30% of terminal azide on the nanowire surface remained unreacted. Temperature-dependent FT-IR measurements revealed that such unreacted residual azides deteriorate the thermal stability of the nanowire molecular layer. To overcome this observed conversion limitation of click reactions on nanostructure surfaces, we considered the steric hindrance around the closely packed SAM reaction points, then experimented with dispersing the azide moiety into a methyl-terminated SAM. The mixed-SAM method significantly improved the azide conversion rate to almost 100%. This reaction method enables the construction of spatially patterned molecular surface modifications on metal oxide nanowire arrays without detrimental unreacted azide groups.

Synthesis of Monodispersedly Sized ZnO Nanowires from Randomly Sized Seeds.

We demonstrate the facile, rational synthesis of monodispersedly sized zinc oxide (ZnO) nanowires from randomly sized seeds by hydrothermal growth. Uniformly shaped nanowire tips constructed in ammonia-dominated alkaline conditions serve as a foundation for the subsequent formation of the monodisperse nanowires. By precisely controlling the sharp tip formation and the nucleation, our method substantially narrows the distribution of ZnO nanowire diameters from σ = 13.5 nm down to σ = 1.3 nm and controls their diameter by a completely bottom-up method, even initiating from randomly sized seeds. The proposed concept of sharp tip based monodisperse nanowires growth can be applied to the growth of diverse metal oxide nanowires and thus paves the way for bottom-up grown metal oxide nanowires-integrated nanodevices with a reliable performance.

Unusual Oxygen Partial Pressure Dependence of Electrical Transport of Single-Crystalline Metal Oxide Nanowires Grown by the Vapor-Liquid-Solid Process.

In general, the electrical conductivities of n-type semiconducting metal oxide nanostructures increase with the decrease in the oxygen partial pressure during crystal growth owing to the increased number of crystal imperfections including oxygen vacancies. In this paper, we report an unusual oxygen partial pressure dependence of the electrical conductivity of single-crystalline SnO2 nanowires grown by a vapor-liquid-solid (VLS) process. The electrical conductivity of a single SnO2 nanowire, measured using the four-probe method, substantially decreases by 2 orders of magnitude when the oxygen partial pressure for the crystal growth is reduced from 10-3 to 10-4 Pa. This contradicts the conventional trend of n-type SnO2 semiconductors. Spatially resolved single-nanowire electrical transport measurements, microstructure analysis, plane-view electron energy-loss spectroscopy, and molecular dynamics simulations reveal that the observed unusual oxygen partial pressure dependence of the electrical transport is attributed to the intrinsic differences between the two crystal growth interfaces (LS and VS interfaces) in the critical nucleation of the crystal growth and impurity incorporation probability as a function of the oxygen partial pressure. The impurity incorporation probability at the LS interface is always lower than that at the VS interface, even under reduced oxygen partial pressures.

Rational Method of Monitoring Molecular Transformations on Metal-Oxide Nanowire Surfaces.

Metal-oxide nanowires have demonstrated excellent capability in the electrical detection of various molecules based on their material robustness in liquid and air environments. Although the surface structure of the nanowires essentially determines their interaction with adsorbed molecules, understanding the correlation between an oxide nanowire surface and an adsorbed molecule is still a major challenge. Herein, we propose a rational methodology to obtain this information for low-density molecules adsorbed on metal oxide nanowire surfaces by employing infrared p-polarized multiple-angle incidence resolution spectroscopy and temperature-programmed desorption/gas chromatography-mass spectrometry. As a model system, we studied the surface chemical transformation of an aldehyde (nonanal, a cancer biomarker in breath) on single-crystalline ZnO nanowires. We found that a slight surface reconstruction, induced by the thermal pretreatment, determines the surface chemical reactivity of nonanal. The present results show that the observed surface reaction trend can be interpreted in terms of the density of Zn ions exposed on the nanowire surface and of their corresponding spatial arrangement on the surface, which promotes the reaction between neighboring adsorbed molecules. The proposed methodology will support a better understanding of complex molecular transformations on various nanostructured metal-oxide surfaces.

True Vapor-Liquid-Solid Process Suppresses Unintentional Carrier Doping of Single Crystalline Metal Oxide Nanowires.

Single crystalline nanowires composed of semiconducting metal oxides formed via a vapor-liquid-solid (VLS) process exhibit an electrical conductivity even without an intentional carrier doping, although these stoichiometric metal oxides are ideally insulators. Suppressing this unintentional doping effect has been a challenging issue not only for metal oxide nanowires but also for various nanostructured metal oxides toward their semiconductor applications. Here we demonstrate that a pure VLS crystal growth, which occurs only at liquid-solid (LS) interface, substantially suppresses an unintentional doping of single crystalline SnO2 nanowires. By strictly tailoring the crystal growth interface of VLS process, we found the gigantic difference of electrical conduction (up to 7 orders of magnitude) between nanowires formed only at LS interface and those formed at both LS and vapor-solid (VS) interfaces. On the basis of investigations with spatially resolved single nanowire electrical measurements, plane-view electron energy-loss spectroscopy, and molecular dynamics simulations, we reveal the gigantic suppression of unintentional carrier doping only for the crystal grown at LS interface due to the higher annealing effect at LS interface compared with that grown at VS interface. These implications will be a foundation to design the semiconducting properties of various nanostructured metal oxides.

Substantial Expansion of Detectable Size Range in Ionic Current Sensing through Pores by Using a Microfluidic Bridge Circuit.

Measuring ionic currents passing through nano- or micropores has shown great promise for the electrical discrimination of various biomolecules, cells, bacteria, and viruses. However, conventional measurements have shown there is an inherent limitation to the detectable particle volume (1% of the pore volume), which critically hinders applications to real mixtures of biomolecule samples with a wide size range of suspended particles. Here we propose a rational methodology that can detect samples with the detectable particle volume of 0.01% of the pore volume by measuring a transient current generated from the potential differences in a microfluidic bridge circuit. Our method substantially suppresses the background ionic current from the µA level to the pA level, which essentially lowers the detectable particle volume limit even for relatively large pore structures. Indeed, utilizing a microscale long pore structure (volume of 5.6 × 104 aL; height and width of 2.0 × 2.0 µm; length of 14 µm), we successfully detected various samples including polystyrene nanoparticles (volume: 4 aL), bacteria, cancer cells, and DNA molecules. Our method will expand the applicability of ionic current sensing systems for various mixed biomolecule samples with a wide size range, which have been difficult to measure by previously existing pore technologies.

Rational Concept for Reducing Growth Temperature in Vapor-Liquid-Solid Process of Metal Oxide Nanowires.

Vapor-liquid-solid (VLS) growth process of single crystalline metal oxide nanowires has proven the excellent ability to tailor the nanostructures. However, the VLS process of metal oxides in general requires relatively high growth temperatures, which essentially limits the application range. Here we propose a rational concept to reduce the growth temperature in VLS growth process of various metal oxide nanowires. Molecular dynamics (MD) simulation theoretically predicts that it is possible to reduce the growth temperature in VLS process of metal oxide nanowires by precisely controlling the vapor flux. This concept is based on the temperature dependent "material flux window" that the appropriate vapor flux for VLS process of nanowire growth decreases with decreasing the growth temperature. Experimentally, we found the applicability of this concept for reducing the growth temperature of VLS processes for various metal oxides including MgO, SnO2, and ZnO. In addition, we show the successful applications of this concept to VLS nanowire growths of metal oxides onto tin-doped indium oxide (ITO) glass and polyimide (PI) substrates, which require relatively low growth temperatures.

Identifying DNA methylation in a nanochannel.

DNA methylation is a stable epigenetic modification, which is well known to be involved in gene expression regulation. In general, however, analyzing DNA methylation requires rather time consuming processes (24-96 h) via DNA replication and protein modification. Here we demonstrate a methodology to analyze DNA methylation at a single DNA molecule level without any protein modifications by measuring the contracted length and relaxation time of DNA within a nanochannel. Our methodology is based on the fact that methylation makes DNA molecules stiffer, resulting in a longer contracted length and a longer relaxation time (a slower contraction rate). The present methodology offers a promising way to identify DNA methylation without any protein modification at a single DNA molecule level within 2 h.

Rational Concept for Designing Vapor-Liquid-Solid Growth of Single Crystalline Metal Oxide Nanowires.

Metal oxide nanowires hold great promise for various device applications due to their unique and robust physical properties in air and/or water and also due to their abundance on Earth. Vapor-liquid-solid (VLS) growth of metal oxide nanowires offers the high controllability of their diameters and spatial positions. In addition, VLS growth has applicability to axial and/or radial heterostructures, which are not attainable by other nanowire growth methods. However, material species available for the VLS growth of metal oxide nanowires are substantially limited even though the variety of material species, which has fascinating physical properties, is the most interesting feature of metal oxides. Here we demonstrate a rational design for the VLS growth of various metal oxide nanowires, based on the "material flux window". This material flux window describes the concept of VLS nanowire growth within a limited material flux range, where nucleation preferentially occurs only at a liquid-solid interface. Although the material flux was previously thought to affect primarily the growth rate, we experimentally and theoretically demonstrate that the material flux is the important experimental variable for the VLS growth of metal oxide nanowires. On the basis of the material flux window concept, we discover novel metal oxide nanowires, composed of MnO, CaO, Sm2O3, NiO, and Eu2O3, which were previously impossible to form via the VLS route. The newly grown NiO nanowires exhibited stable memristive properties superior to conventional polycrystalline devices due to the single crystallinity. Thus, this VLS design route offers a useful guideline for the discovery of single crystalline nanowires that are composed of functional metal oxide materials.

Isopentenyl pyrophosphate secreted from Zoledronate-stimulated myeloma cells, activates the chemotaxis of γδT cells.

γδT cell receptor (TCR)-positive T cells, which control the innate immune system, display anti-tumor immunity as well as other non-immune-mediated anti-cancer effects. γδT cells expanded ex vivo by nitrogen-containing bisphosphonate (N-BP) treatment can kill tumor cells. N-BP inhibits farnesyl pyrophosphate synthase in the mevalonate pathway, resulting in the accumulation of isopentenyl pyrophosphate (IPP), which is a stimulatory antigen for γδT cells. We have previously observed that as they get closer, migrating γδT cells increase in speed toward target multiple myeloma (MM) cells. In the present study, we investigated the γδT cell chemotactic factors involving using a micro total analysis system-based microfluidic cellular analysis device. The addition of supernatant from RPMI8226 MM cells treated with the N-BP zoledronic acid (ZOL) or the addition of IPP to the device induced chemotaxis of γδT cells and increased the speed of migration compared to controls. Analysis of the ZOL-treated RPMI8226 cell supernatant revealed that it contained IPP secreted in a ZOL-dose-dependent manner. These observations indicate that IPP activates the chemotaxis of γδT cells toward target MM cells treated with ZOL.

Modulation of thermoelectric power factor via radial dopant inhomogeneity in B-doped Si nanowires.

We demonstrate a modulation of thermoelectric power factor via a radial dopant inhomogeneity in B-doped Si nanowires. These nanowires grown via vapor-liquid-solid (VLS) method were naturally composed of a heavily doped outer shell layer and a lightly doped inner core. The thermopower measurements for a single nanowire demonstrated that the power factor values were higher than those of homogeneously B-doped Si nanowires. The field effect measurements revealed the enhancement of hole mobility for these VLS grown B-doped Si nanowires due to the modulation doping effect. This mobility enhancement increases overall electrical conductivity of nanowires without decreasing the Seebeck coefficient value, resulting in the increase of thermoelectric power factor. In addition, we found that tailoring the surface dopant distribution by introducing surface δ-doping can further increase the power factor value. Thus, intentionally tailoring radial dopant inhomogeneity promises a way to modulate the thermoelectric power factor of semiconductor nanowires.

Efficient hepatocyte differentiation of primary human hepatocyte-derived organoids using three dimensional nanofibers (HYDROX) and their possible application in hepatotoxicity research.

Human liver organoids are in vitro three dimensionally (3D) cultured cells that have a bipotent stem cell phenotype. Translational research of human liver organoids for drug discovery has been limited by the challenge of their low hepatic function compared to primary human hepatocytes (PHHs). Various attempts have been made to develop functional hepatocyte-like cells from human liver organoids. However, none have achieved the same level of hepatic functions as PHHs. We here attempted to culture human liver organoids established from cryopreserved PHHs (PHH-derived organoids), using HYDROX, a chemically defined 3D nanofiber. While the proliferative capacity of PHH-derived organoids was lost by HYDROX-culture, the gene expression levels of drug-metabolizing enzymes were significantly improved. Enzymatic activities of cytochrome P450 3A4 (CYP3A4), CYP2C19, and CYP1A2 in HYDROX-cultured PHH-derived organoids (Org-HYDROX) were comparable to those in PHHs. When treated with hepatotoxic drugs such as troglitazone, amiodarone and acetaminophen, Org-HYDROX showed similar cell viability to PHHs, suggesting that Org-HYDROX could be applied to drug-induced hepatotoxicity tests. Furthermore, Org-HYDROX maintained its functions for up to 35 days and could be applied to chronic drug-induced hepatotoxicity tests using fialuridine. Our findings demonstrated that HYDROX could possibly be a novel biomaterial for differentiating human liver organoids towards hepatocytes applicable to pharmaceutical research.

Impact of preferential indium nucleation on electrical conductivity of vapor-liquid-solid grown indium-tin oxide nanowires.

Highly conductive and transparent indium-tin oxide (ITO) single-crystalline nanowires, formed by the vapor-liquid-solid (VLS) method, hold great promise for various nanoscale device applications. However, increasing an electrical conductivity of VLS grown ITO nanowires is still a challenging issue due to the intrinsic difficulty in controlling complex material transports of the VLS process. Here, we demonstrate a crucial role of preferential indium nucleation on the electrical conductivity of VLS grown ITO nanowires using gold catalysts. In spite of the fact that the vapor pressure of tin is lower than that of indium, we found that the indium concentration within the nanowires was always higher than the nominal composition. The VLS growth of ITO through gold catalysts significantly differs from ITO film formations due to the emergence of preferential indium nucleation only at a liquid-solid interface. Furthermore, we demonstrate that the averaged resistivity of ITO nanowires can be decreased down to 2.1 × 10(-4) Ω cm, which is the lowest compared with values previously reported, via intentionally increasing the tin concentration within the nanowires.

Prominent thermodynamical interaction with surroundings on nanoscale memristive switching of metal oxides.

This study demonstrates the effect of surroundings on a memristive switching at nanoscale by utilizing an open top planar-type device. NiO(x) and CoO(x) planar-type devices have exhibited a memristive behavior under atmospheric pressure, whereas TiO(2-x) planar-type devices did not show a memristive switching even under the same surroundings. A memristive behavior of TiO(2-x) planar-type devices has emerged when reducing an ambient pressure and/or employing a SiO(2) passivation layer. These results reveal that a thermodynamical interaction with surroundings critically determines the occurrence of memristive switching via varying a stability of nonstoichiometry. Since this effect tends to be more significant for smaller devices with larger specific surface area, tailoring the surrounding effect by an appropriate passivation will be essential for high density devices.

Glow-type conversion and characterization of a minimal luciferase via mutational analyses.

Luciferases are widely used as reporter proteins in various fields. Recently, we developed a minimal bright luciferase, picALuc, via partial deletion of the artificial luciferase (ALuc) derived from copepods luciferases. However, the structures of copepod luciferases in the substrate-bound state remain unknown. Moreover, as suggested by structural modeling, picALuc has a larger active site cavity, unlike that in other copepod luciferases. Here, to explore the bioluminescence mechanism of picALuc and its luminescence properties, we conducted multiple mutational analyses, and identified residues and regions important for catalysis and bioluminescence. Mutations of residues likely involved in catalysis (S33, H34, and D55) markedly reduced bioluminescence, whereas that of residue (E50) (near the substrate in the structural model) enhanced luminescence intensity. Furthermore, deletion mutants (Δ70-Δ78) in the loop region (around I73) exhibited longer luminescence lifetimes (~ 30 min) and were reactivated multiple times upon re-addition of the substrate. Due to the high thermostability of picALuc, one of its representative mutant (Δ74), was able to be reused, that is, luminescence recycling, for day-scale time at room temperature. These findings provide important insights into picALuc bioluminescence mechanism and copepod luciferases and may help with sustained observations in a variety of applications.

Improving bioluminescence of a minimal luciferase by adding a charged oligopeptide: picALuc2.0.

Luciferases are widely used as reporter proteins in diverse fields from basic biology to medical and environmental researches. Development of luciferase applications for reporter proteins requires small size without target inhibition, appropriate genomic insertion for high expression level, and bright emission for detection sensitivity. We previously developed the minimal luciferase picALuc, but its luminescence was still dim compared to other bright luciferases in terms of expression in Escherichia coli. In this study, diverse additions of oligopeptides with charged residues (eight amino acids in total) to the C-terminus of picALuc enhanced luminescence by up to approximately 50-fold, that is, enhanced enzymatic activity. Moreover, these high luminescence activities were achieved in bacterial and mammalian expression, suggesting their further applicability in many expression systems. The finding in this study that the simple addition of oligopeptides with charged residues (or charge engineering of this kind) enhances enzymatic activity may be applied to a wide variety of enzymatic reactions and protein functions.

Interfacial Molecular Compatibility for Programming Organic-Metal Oxide Superlattices.

Artificially programming a sequence of organic-metal oxide multilayers (superlattices) by using atomic layer deposition (ALD) is a fascinating and challenging issue in material chemistry. However, the complex chemical reactions between ALD precursors and organic layer surfaces have limited their applications for various material combinations. Here, we demonstrate the impact of interfacial molecular compatibility on the formation of organic-metal oxide superlattices using ALD. The effects of both organic and inorganic compositions on the metal oxide layer formation processes onto self-assembled monolayers (SAM) were examined by using scanning transmission electron microscopy, in situ quartz crystal microbalance measurements, and Fourier-transformed infrared spectroscopy. These series of experiments reveal that the terminal group of organic SAM molecules must satisfy two conflicting requirements, the first of which is to promptly react with ALD precursors and the second is not to bind strongly to the bottom metal oxide layers to avoid undesired SAM conformations. OH-terminated phosphate aliphatic molecules, which we have synthesized, were identified as one of the best candidates for such a purpose. Molecular compatibility between metal oxide precursors and the -OHs must be properly considered to form superlattices. In addition, it is also important to form densely packed and all-trans-like SAMs to maximize the surface density of reactive -OHs on the SAMs. Based on these design strategies for organic-metal oxide superlattices, we have successfully fabricated various superlattices composed of metal oxides (Al-, Hf-, Mg-, Sn-, Ti-, and Zr oxides) and their multilayered structures.

Rational Strategy for Space-Confined Atomic Layer Deposition.

Atomic layer deposition (ALD) offers excellent controllability of spatial uniformity, film thickness at the Angstrom level, and film composition even for high-aspect-ratio nanostructured surfaces, which are rarely attainable by other conventional deposition methodologies. Although ALD has been successfully applied to various substrates under open-top circumstances, the applicability of ALD to confined spaces has been limited because of the inherent difficulty of supplying precursors into confined spaces. Here, we propose a rational methodology to apply ALD growths to confined spaces (meter-long microtubes with an aspect ratio of up to 10 000). The ALD system, which can generate differential pressures to confined spaces, was newly developed. By using this ALD system, it is possible to deposit TiOx layers onto the inner surface of capillary tubes with a length of 1000 mm and an inner diameter of 100 µm with spatial deposition uniformity. Furthermore, we show the superior thermal and chemical robustness of TiOx-coated capillary microtubes for molecular separations when compared to conventional molecule-coated capillary microtubes. Thus, the present rational strategy of space-confined ALD offers a useful approach to design the chemical and physical properties of the inner surfaces of various confined spaces.