Zn2+-Depletion Enhances Lysosome Fission in Cultured Rat Embryonic Cortical Neurons Revealed by a Modified Epifluorescence Microscopic Technique.

Lysosomes are integration hubs for several signaling pathways, such as autophagy and endocytosis, and also crucial stores of ions, including Zn2+. Lysosomal dysfunction caused by changes in their morphology by fusion and fission processes can result in several pathological disorders. However, the role of Zn2+ in modulating the morphology of lysosomes is unclear. The resolution of conventional epifluorescence microscopy restricts accurate observation of morphological changes of subcellular fluorescence punctum. In this study, we used a modified epifluorescence microscopy to identify the center of a punctum from a series of z-stack images and calculate the morphological changes. We stained primary cultured rat embryonic cortical neurons with FluoZin3, a Zn2+-sensitive fluorescent dye, and Lysotracker, a lysosome-specific marker, to visualize the distribution of Zn2+-enriched vesicles and lysosomes, respectively. Our results revealed that treating neurons with N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine, a cell-permeable Zn2+ chelator, shrank Zn2+-enriched vesicles and lysosomes by up to 25% in an hour. Pretreating the neurons with YM201636, a blocker of lysosome fission, could suppress this shrinkage. These results demonstrate the usefulness of the modified epifluorescence microscopy for investigating the homeostasis of intracellular organelles and related disorders.

Intracellular Delivery of Luciferase with Fluorescent Nanodiamonds for Dual-Modality Imaging of Human Stem Cells.

Delivering functional proteins (such as enzymes) into cells is important in various biological studies and is often accomplished indirectly by transfection with DNA or mRNA encoding recombinant proteins. However, the transfection efficiency of conventional plasmid methods is low for primary cells, which are crucial sources of cell therapy. Here, we present a new platform based on the use of fluorescent nanodiamond (FND) as a biocompatible nanocarrier to enable rapid, effective, and homogeneous labeling of human mesenchymal stem cells (MSCs) with luciferase for multiplex assays and ultrasensitive detection. More than 100 pg of FND and 100 million copies of firefly luciferase can be delivered into each MSC through endocytosis. Moreover, these endocytic luciferase molecules are catalytically active for hours, allowing the cells to be imaged and tracked in vitro as well as in vivo by both fluorescence and bioluminescence imaging. Our results demonstrate that luciferase-conjugated FNDs are useful as multifunctional labels of human stem cells for diverse theranostic applications.

Tuning Rashba Spin-Orbit Coupling in Gated Multilayer InSe.

Manipulating the electron spin with the aid of spin-orbit coupling (SOC) is an indispensable element of spintronics. Electrostatically gating a material with strong SOC results in an effective magnetic field which can in turn be used to govern the electron spin. In this work, we report the existence and electrostatic tunability of Rashba SOC in multilayer InSe. We observed a gate-voltage-tuned crossover from weak localization (WL) to weak antilocalization (WAL) effect in quantum transport studies of InSe, which suggests an increasing SOC strength. Quantitative analyses of magneto-transport studies and energy band diagram calculations provide strong evidence for the predominance of Rashba SOC in electrostatically gated InSe. Furthermore, we attribute the tendency of the SOC strength to saturate at high gate voltages to the increased electronic density of states-induced saturation of the electric field experienced by the electrons in the InSe layer. This explanation of nonlinear gate voltage control of Rashba SOC can be generalized to other electrostatically gated semiconductor nanomaterials in which a similar tendency of spin-orbit length saturation was observed (e.g., nanowire field effect transistors), and is thus of broad implications in spintronics. Identifying and controlling the Rashba SOC in InSe may serve pivotally in devising III-VI semiconductor-based spintronic devices in the future.

The Extracellular Zn2+ Concentration Surrounding Excited Neurons Is High Enough to Bind Amyloid-ß Revealed by a Nanowire Transistor.

The Zn2+ stored in the secretory vesicles of glutamatergic neurons is coreleased with glutamate upon stimulation, resulting in the elevation of extracellular Zn2+ concentration (CZn2+ex). This elevation of CZn2+ex regulates the neurotransmission and facilitates the fibrilization of amyloid-ß (Aß). However, the exact CZn2+ex surrounding neurons under (patho)physiological conditions is not clear and the connection between CZn2+ex and the Aß fibrilization remains obscure. Here, a silicon nanowire field-effect transistor (SiNW-FET) with the Zn2+ -sensitive fluorophore, FluoZin-3 (FZ-3), to quantify the CZn2+ex in real time is modified. This FZ-3/SiNW-FET device has a dissociation constant of ≈12 × 10-9 m against Zn2+ . By placing a coverslip seeded with cultured embryonic cortical neurons atop an FZ-3/SiNW-FET, the CZn2+ex elevated to ≈110 × 10-9 m upon stimulation with α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA). Blockers against the AMPA receptor or exocytosis greatly suppress this elevation, indicating that the Zn2+ stored in the synaptic vesicles is the major source responsible for this elevation of CZn2+ex. In addition, a SiNW-FET modified with Aß could bind Zn2+ with a dissociation constant of ≈633 × 10-9 m and respond to the Zn2+ released from AMPA-stimulated neurons. Therefore, the CZn2+ex can reach a level high enough to bind Aß and the Zn2+ homeostasis can be a therapeutic strategy to prevent neurodegeneration.

Detection of electrically neutral and nonpolar molecules in ionic solutions using silicon nanowires.

We report on a technique that can extend the use of nanowire sensors to the detection of interactions involving nonpolar and neutral molecules in an ionic solution environment. This technique makes use of the fact that molecular interactions result in a change in the permittivity of the molecules involved. For the interactions taking place at the surface of nanowires, this permittivity change can be determined from the analysis of the measured complex impedance of the nanowire. To demonstrate this technique, histidine was detected using different charge polarities controlled by the pH value of the solution. This included the detection of electrically neutral histidine at a sensitivity of 1 pM. Furthermore, it is shown that nonpolar molecules, such as hexane, can also be detected. The technique is applicable to the use of nanowires with and without a surface-insulating oxide. We show that information about the changes in amplitude and the phase of the complex impedance reveals the fundamental characteristics of the molecular interactions, including the molecular field and the permittivity.

Differential Releases of Dopamine and Neuropeptide Y from Histamine-Stimulated PC12 Cells Detected by an Aptamer-Modified Nanowire Transistor.

Silicon nanowire field-effect transistors modified with specific aptamers can directly detect the minute dopamine and neuropeptide Y released from cells. The binding of these molecules to the aptamers results in a conductance change of the transistor biosensor and illustrates the differential releasing mechanisms of these molecules stored in various vesicle pools.

Intrinsic Electron Mobility Exceeding 10³ cm²/(V s) in Multilayer InSe FETs.

Graphene-like two-dimensional (2D) materials not only are interesting for their exotic electronic structure and fundamental electronic transport or optical properties but also hold promises for device miniaturization down to atomic thickness. As one material belonging to this category, InSe, a III-VI semiconductor, not only is a promising candidate for optoelectronic devices but also has potential for ultrathin field effect transistor (FET) with high mobility transport. In this work, various substrates such as PMMA, bare silicon oxide, passivated silicon oxide, and silicon nitride were used to fabricate multilayer InSe FET devices. Through back gating and Hall measurement in four-probe configuration, the device's field effect mobility and intrinsic Hall mobility were extracted at various temperatures to study the material's intrinsic transport behavior and the effect of dielectric substrate. The sample's field effect and Hall mobilities over the range of 20-300 K fall in the range of 0.1-2.0 × 10(3) cm(2)/(V s), which are comparable or better than the state of the art FETs made of widely studied 2D transition metal dichalcogenides.

Tracking and Finding Slow-Proliferating/Quiescent Cancer Stem Cells with Fluorescent Nanodiamonds.

Quiescent cancer stem cells (CSCs) have long been considered to be a source of tumor initiation. However, identification and isolation of these cells have been hampered by the fact that commonly used fluorescent markers are not sufficiently stable, both chemically and photophysically, to allow tracking over an extended period of time. Here, it is shown that fluorescent nanodiamonds (FNDs) are well suited for this application. Genotoxicity tests of FNDs with comet and micronucleus assays for human fibroblasts and breast cancer cells indicate that the nanoparticles neither cause DNA damage nor impair cell growth. Using AS-B145-1R breast cancer cells as the model cell line for CSC, it is found that the FND labeling outperforms 5-ethynyl-2'-deoxyuridine (EdU) and carboxyfluorescein diacetate succinimidyl ester (CFSE) in regards to its long-term tracking capability (>20 d). Moreover, through a quantification of their stem cell activity by measuring mammosphere-forming efficiencies (MFEs) and self-renewal rates, the FND-positive cells are identified to have an MFE twice as high as that of the FND-negative cells isolated from the same dissociated mammospheres. Thus, the nanoparticle-based labeling technique provides an effective new tool for tracking and finding slow-proliferating/quiescent CSCs in cancer research.

High performance and bendable few-layered InSe photodetectors with broad spectral response.

Two-dimensional crystals with a wealth of exotic dimensional-dependent properties are promising candidates for next-generation ultrathin and flexible optoelectronic devices. For the first time, we demonstrate that few-layered InSe photodetectors, fabricated on both a rigid SiO2/Si substrate and a flexible polyethylene terephthalate (PET) film, are capable of conducting broadband photodetection from the visible to near-infrared region (450-785 nm) with high photoresponsivities of up to 12.3 AW(-1) at 450 nm (on SiO2/Si) and 3.9 AW(-1) at 633 nm (on PET). These photoresponsivities are superior to those of other recently reported two-dimensional (2D) crystal-based (graphene, MoS2, GaS, and GaSe) photodetectors. The InSe devices fabricated on rigid SiO2/Si substrates possess a response time of ∼50 ms and exhibit long-term stability in photoswitching. These InSe devices can also operate on a flexible substrate with or without bending and reveal comparable performance to those devices on SiO2/Si. With these excellent optoelectronic merits, we envision that the nanoscale InSe layers will not only find applications in flexible optoelectronics but also act as an active component to configure versatile 2D heterostructure devices.

Photoelectrochemical activity on Ga-polar and N-polar GaN surfaces for energy conversion.

Hydrogen generation through direct photoelectrolysis of water was studied using photoelectrochemical cells made of different facets of free-standing polar GaN system. To build the fundamental understanding at the differences of surface photochemistry afforded by the GaN {0001}and {000-1}polar surfaces, we correlated the relationship between the surface structure and photoelectrochemical performance on the different polar facets. The photoelectrochemical measurements clearly revealed that the Ga-polar surface had a more negative onset potential relative to the N-polar surface due to the much negative flat-band potential. At more positive applied voltages, however, the N-polar surface yielded much higher photocurrent with conversion efficiency of 0.61% compared to that of 0.55% by using the Ga-polar surface. The reason could be attributed to the variation in the band structure of the different polar facets via Mott-Schottky analyses. Based on this work, understanding the facet effect on photoelectrochemical activity can provide a blueprint for the design of materials in solar hydrogen applications.

Advances in nanowire transistors for biological analysis and cellular investigation.

Electrical biosensors based on silicon nanowire field-effect transistors (SiNW-FETs) have attracted enormous interest in the biosensing field. SiNW-FETs have proven to be significant and efficient in detecting diverse biomolecular species with the advantages of high probing sensitivity, target selectivity, real-time recording and label-free detection. In recent years, significant advances in biosensors have been achieved, particularly for cellular investigation and biomedical diagnosis. In this critical review, we will report on the latest developments in biosensing with SiNW-FETs and discuss recent advancements in the innovative designs of SiNW-FET devices. This critical review introduces the basic instrumental setup and working principle of SiNW-FETs. Technical approaches that attempted to enhance the detection sensitivity and target selectivity of SiNW-FET sensors are discussed. In terms of applications, we review the recent achievements with SiNW-FET biosensors for the investigations of protein-protein interaction, DNA/RNA/PNA hybridization, virus detection, cellular recording, biological kinetics, and clinical diagnosis. In addition, the novel architecture designs of the SiNW-FET devices are highlighted in studies of live neuron cells, electrophysiological measurements and other signal transduction pathways. Despite these remarkable achievements, certain improvements remain necessary in the device performance and clinical applications of FET-based biosensors; thus, several prospects about the future development of nanowire transistor-based instruments for biosensing employments are discussed at the end of this review.

Improving nanowire sensing capability by electrical field alignment of surface probing molecules.

We argue that the structure ordering of self-assembled probing molecular monolayers is essential for the reliability and sensitivity of nanowire-based field-effect sensors because it can promote the efficiency for molecular interactions as well as strengthen the molecular dipole field experienced by the nanowires. In the case of monolayers, we showed that structure ordering could be improved by means of electrical field alignment. This technique was then employed to align multilayer complexes for nanowire sensing applications. The sensitivity we achieved for detection of hybridization between 15-base single-strand DNA molecules is 0.1 fM and for alcohol sensors is 0.5 ppm. The reliability was confirmed by repeated tests on chips that contain multiple nanowire sensors.

An ultrasensitive nanowire-transistor biosensor for detecting dopamine release from living PC12 cells under hypoxic stimulation.

Dopamine (DA) is an important neurotransmitter that is involved in neuronal signal transduction and several critical illnesses. However, the concentration of DA is extremely low in patients and is difficult to detect using existing electrochemical biosensors with detection limits typically around nanomolar levels (∼10(-9) M). Here, we developed a nanoelectronic device as a biosensor for ultrasensitive and selective DA detection by modifying DNA-aptamers on a multiple-parallel-connected (MPC) silicon nanowire field-effect transistor (referred to as MPC aptamer/SiNW-FET). Compared with conventional electrochemical methods, the MPC aptamer/SiNW-FET has been demonstrated to improve the limit of DA detection to <10(-11) M and to possess a detection specificity that is able to distinguish DA from other chemical analogues, such as ascorbic acid, catechol, phenethylamine, tyrosine, epinephrine, and norepinephrine. This MPC aptamer/SiNW-FET was also applied to monitor DA release under hypoxic stimulation from living PC12 cells. The real-time recording of the exocytotic DA induced by hypoxia reveals that the increase in intracellular Ca(2+) that is required to trigger DA secretion is dominated by an extracellular Ca(2+) influx, rather than the release of intracellular Ca(2+) stores.

Chemical vapor deposition synthesis and Raman spectroscopic characterization of large-area graphene sheets.

We present a chemical vapor deposition (CVD) method to catalytically synthesize large-area, transferless, single- to few-layer graphene sheets using hexamethyldisilazane (HMDS) on a SiO2/Si substrate as a carbon source and thermally evaporated alternating Ni/Cu/Ni layers as a catalyst. The as-synthesized graphene films were characterized by Raman spectroscopic imaging to identify single- to few-layer sheets. This HMDS-derived graphene layer is continuous over the entire growth substrate, and single- to trilayer mixed sheets can be up to 30 µm in the lateral dimension. With the synthetic CVD method proposed here, graphene can be grown into tailored shapes directly on a SiO2/Si surface through vapor priming of HMDS onto predefined photolithographic patterns. The transparent and conductive HMDS-derived graphene exhibits its potential for widespread electronic and opto-electronic applications.

Label-free detection of protein-protein interactions using a calmodulin-modified nanowire transistor.

In this study, we describe a highly sensitive and reusable silicon nanowire field-effect transistor for the detection of protein-protein interactions. This reusable device was made possible by the reversible association of glutathione S-transferase-tagged calmodulin with a glutathione modified transistor. The calmodulin-modified transistor exhibited selective electrical responses to Ca2+ (> or = 1 microM) and purified cardiac troponin I (approximately 7 nM); the change in conductivity displayed a linear dependence on the concentration of troponin I in a range from 10 nM to 1 microM. These results are consistent with the previously reported concentration range in which the dissociation constant for the troponin I-calmodulin complex was determined. The minimum concentration of Ca2+ required to activate calmodulin was determined to be 1 microM. We have also successfully demonstrated that the N-type Ca2+ channels, expressed by cultured 293T cells, can be recognized specifically by the calmodulin-modified nanowire transistor. This sensitive nanowire transistor can serve as a high-throughput biosensor and can also substitute for immunoprecipitation methods used in the identification of interacting proteins.

Coupling between Pyroelectricity and Built-In Electric Field Enabled Highly Sensitive Infrared Phototransistor Based on InSe/WSe2/P(VDF-TrFE) Heterostructure.

The assorted utilization of infrared detectors induces the demand for more comprehensive and high-performance electronic devices that work at room temperature. The intricacy of the fabrication process with bulk material limits the exploration in this field. However, two-dimensional (2D) materials with a narrow band gap opening aid in infrared (IR) detection relatively, but the photodetection range is narrowed due to the inherent band gap. In this study, we report an unprecedented attempt at the coordinated use of both 2D heterostructure (InSe/WSe2) and the dielectric polymer (poly(vinylidene fluoride-trifluoroethylene), P(VDF-TrFE)) for both visible and IR photodetection in a single device. The remnant polarization due to the ferroelectric effect of the polymer dielectric enhances the photocarrier separation in the visible range, resulting in high photoresponsivity. On the other hand, the pyroelectric effect of the polymer dielectric causes a change in the device current due to the increased temperature induced by the localized heating effect of the IR irradiation, which results in the change of ferroelectric polarization and induces the redistribution of charge carriers. In turn, it changes the built-in electric field, the depletion width, and the band alignment across the p-n heterojunction interface. Consequently, the charge carrier separation and the photosensitivity are therefore enhanced. Through the coupling between pyroelectricity and built-in electric field across the heterojunction, the specific detectivity for the photon energy below the band gap of the constituent 2D materials can reach up to 1011 Jones, which is better than all reported pyroelectric IR detectors. The proposed approach combining the ferroelectric and pyroelectric effects of the dielectric as well as exceptional properties of the 2D heterostructures can spark the design of advanced and not-yet realized optoelectronic devices.

In Situ Spectroelectrochemical Detection of Oxygen Evolution Reaction Intermediates with a Carboxylated Graphene-MnO2 Electrocatalyst.

In electrocatalyst-assisted water splitting, the oxygen evolution reaction (OER) imposes a performance limit due to the formation of different catalyst-bound intermediates and the scaling relationship of their adsorption energies. To break this scaling relationship in OER, a bifunctional mechanism was proposed recently, in which the energetically demanding step of forming the *OOH intermediate, through the attack of a water molecule on the oxo unit (*O, with * representing a reactive metal center), is facilitated by proton transfer to the second catalytic site. This mechanism was supported theoretically but so far by only very few experiments with a proton-transfer agent in basic media. However, active metal-containing catalysts could be destroyed in alkaline media, raising questions on practical applications. To date, this mechanism still lacks a systematic spectroscopic support by observing the short-lived and limited amount of reactive intermediates. Here, we report an operando Raman spectroscopic observation of the OER intermediates in neutral media, for the first time, via a bifunctional mechanism using a carboxylated graphene-MnO2 (represented by Gr-C-MnO2) electrocatalyst. The formation of the Mn-OOH intermediate after the attack of a water molecule on the Mn═O complex is followed by a proton transfer from Mn-OOH to the functionalized carboxylates. The role of the functionalized carboxylates to improve the catalytic efficiency was further confirmed by both pH-dependent and isotope (H/D)-labeling experiments. Furthermore, with a unique strategy of using a hybrid aqueous/nonaqueous electrolyte, the OER was alleviated, allowing sufficient Mn-OH and Mn-OOH intermediates for in situ Raman spectroscopic observation.

Surface potential variations on a silicon nanowire transistor in biomolecular modification and detection.

Using a silicon nanowire field-effect transistor (SiNW-FET) for biomolecule detections, we selected 3-(mercaptopropyl)trimethoxysilane (MPTMS), N-[6-(biotinamido)hexyl]-3(')-(2(')-pyridyldithio) propionamide (biotin-HPDP), and avidin, respectively, as the designated linker, receptor, and target molecules as a study model, where the biotin molecules were modified on the SiNW-FET to act as a receptor for avidin. We applied high-resolution scanning Kelvin probe force microscopy (KPFM) to detect the modified/bound biomolecules by measuring the induced change of the surface potential (ΔΦ(s)) on the SiNW-FET under ambient conditions. After biotin-immobilization and avidin-binding, the ΔΦ(s) on the SiNW-FET characterized by KPFM was demonstrated to correlate to the conductance change inside the SiNW-FET acquired in aqueous solution. The ΔΦ(s) values on the SiNW-FET caused by the same biotin-immobilization and avidin-binding were also measured from drain current versus gate voltage curves (I(d)-V(g)) in both aqueous condition and dried state. For comparison, we also study the ΔΦ(s) values on a Si wafer caused by the same biotin-immobilization and avidin-binding through KPFM and ζ potential measurements. This study has demonstrated that the surface potential measurement on a SiNW-FET by KPFM can be applied as a diagnostic tool that complements the electrical detection with a SiNW-FET sensor. Although the KPFM experiments were carried out under ambient conditions, the measured surface properties of a SiNW-FET are qualitatively valid compared with those obtained by other biosensory techniques performed in liquid environment.

Significant Elevation in Potassium Concentration Surrounding Stimulated Excitable Cells Revealed by an Aptamer-Modified Nanowire Transistor.

Recording ion fluctuations surrounding biological cells with a nanoelectronic device offers seamless integration of nanotechnology into living organisms and is essential for understanding cellular activities. The concentration of potassium ion in the extracellular fluid (CK+ex) is a critical determinant of cell membrane potential and must be maintained within an appropriate range. Alteration in CK+ex can affect neuronal excitability, induce heart arrhythmias, and even trigger seizure-like reactions in the brain. Therefore, monitoring local fluctuations in real time provides an early diagnosis of the occurrence of the K+-induced pathophysiological responses. Here, we modified the surface of a silicon nanowire field-effect transistor (SiNW-FET) with K+-specific DNA-aptamers (AptK+) to monitor the real-time variations of CK+ex in primary cultured rat embryonic cortical neurons or human embryonic stem cell-derived cardiomyocytes. The binding affinity of AptK+ to K+, determined by measuring the dissociation constant of the AptK+-K+ complex (Kd = 10.1 ± 0.9 mM), is at least 38-fold higher than other ions (e.g., Na+, Ca2+, and Mg2+). By placing cultured cortical neurons over an AptK+/SiNW-FET device, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) stimulation raised the CK+ex dose-dependently to 16 mM when AMPA concentration was >10 µM; this elevation could be significantly suppressed by an AMPA receptor antagonist, 6,7-dinitroquinoxaline-2,3-dione. Likewise, the stimulation of isoproterenol to cardiomyocytes raised the CK+ex to 6-8 mM, with a concomitant increase in the beating rate. This study utilizing a robust nanobiosensor to detect real-time ion fluctuations surrounding excitable cells underlies the importance of ion homeostasis and offers the feasibility of developing an implant device for real-time monitoring.

A Bi-Anti-Ambipolar Field Effect Transistor.

Multistate logic is recognized as a promising approach to increase the device density of microelectronics, but current approaches are offset by limited performance and large circuit complexity. We here demonstrate a route toward increased integration density that is enabled by a mechanically tunable device concept. Bi-anti-ambipolar transistors (bi-AATs) exhibit two distinct peaks in their transconductance and can be realized by a single 2D-material heterojunction-based solid-state device. Dynamic deformation of the device reveals the co-occurrence of two conduction pathways to be the origin of this previously unobserved behavior. Initially, carrier conduction proceeds through the junction edge, but illumination and application of strain can increase the recombination rate in the junction sufficiently to support an alternative carrier conduction path through the junction area. Optical characterization reveals a tunable emission pattern and increased optoelectronic responsivity that corroborates our model. Strain control permits the optimization of the conduction efficiency through both pathways and can be employed in quaternary inverters for future multilogic applications.