Preparing Better Samples for Cryo-Electron Microscopy: Biochemical Challenges Do Not End with Isolation and Purification.

The preparation of extremely thin samples, which are required for high-resolution electron microscopy, poses extreme risk of damaging biological macromolecules due to interactions with the air-water interface. Although the rapid increase in the number of published structures initially gave little indication that this was a problem, the search for methods that substantially mitigate this hazard is now intensifying. The two main approaches under investigation are (a) immobilizing particles onto structure-friendly support films and (b) reducing the length of time during which such interactions may occur. While there is little possibility of outrunning diffusion to the interface, intentional passivation of the interface may slow the process of adsorption and denaturation. In addition, growing attention is being given to gaining more effective control of the thickness of the sample prior to vitrification.

Control of proton transport and hydrogenation in double-gated graphene.

The basal plane of graphene can function as a selective barrier that is permeable to protons1,2 but impermeable to all ions3,4 and gases5,6, stimulating its use in applications such as membranes1,2,7,8, catalysis9,10 and isotope separation11,12. Protons can chemically adsorb on graphene and hydrogenate it13,14, inducing a conductor-insulator transition that has been explored intensively in graphene electronic devices13-17. However, both processes face energy barriers1,12,18 and various strategies have been proposed to accelerate proton transport, for example by introducing vacancies4,7,8, incorporating catalytic metals1,19 or chemically functionalizing the lattice18,20. But these techniques can compromise other properties, such as ion selectivity21,22 or mechanical stability23. Here we show that independent control of the electric field, E, at around 1 V nm-1, and charge-carrier density, n, at around 1 × 1014 cm-2, in double-gated graphene allows the decoupling of proton transport from lattice hydrogenation and can thereby accelerate proton transport such that it approaches the limiting electrolyte current for our devices. Proton transport and hydrogenation can be driven selectively with precision and robustness, enabling proton-based logic and memory graphene devices that have on-off ratios spanning orders of magnitude. Our results show that field effects can accelerate and decouple electrochemical processes in double-gated 2D crystals and demonstrate the possibility of mapping such processes as a function of E and n, which is a new technique for the study of 2D electrode-electrolyte interfaces.

Terahertz phonon engineering with van der Waals heterostructures.

Phonon engineering at gigahertz frequencies forms the foundation of microwave acoustic filters1, acousto-optic modulators2 and quantum transducers3,4. Terahertz phonon engineering could lead to acoustic filters and modulators at higher bandwidth and speed, as well as quantum circuits operating at higher temperatures. Despite their potential, methods for engineering terahertz phonons have been limited due to the challenges of achieving the required material control at subnanometre precision and efficient phonon coupling at terahertz frequencies. Here we demonstrate the efficient generation, detection and manipulation of terahertz phonons through precise integration of atomically thin layers in van der Waals heterostructures. We used few-layer graphene as an ultrabroadband phonon transducer that converts femtosecond near-infrared pulses to acoustic-phonon pulses with spectral content up to 3 THz. A monolayer WSe2 is used as a sensor. The high-fidelity readout was enabled by the exciton-phonon coupling and strong light-matter interactions. By combining these capabilities in a single heterostructure and detecting responses to incident mechanical waves, we performed terahertz phononic spectroscopy. Using this platform, we demonstrate high-Q terahertz phononic cavities and show that a WSe2 monolayer embedded in hexagonal boron nitride can efficiently block the transmission of terahertz phonons. By comparing our measurements to a nanomechanical model, we obtained the force constants at the heterointerfaces. Our results could enable terahertz phononic metamaterials for ultrabroadband acoustic filters and modulators and could open new routes for thermal engineering.

Intelligent infrared sensing enabled by tunable moiré quantum geometry.

Quantum geometric properties of Bloch wave functions in solids, that is, Berry curvature and the quantum metric, are known to significantly influence the ground- and excited-state behaviour of electrons1-5. The bulk photovoltaic effect (BPVE), a nonlinear phenomenon depending on the polarization of excitation light, is largely governed by the quantum geometric properties in optical transitions6-10. Infrared BPVE has yet to be observed in graphene or moiré systems, although exciting strongly correlated phenomena related to quantum geometry have been reported in this emergent platform11-14. Here we report the observation of tunable mid-infrared BPVE at 5 µm and 7.7 µm in twisted double bilayer graphene (TDBG), arising from the moiré-induced strong symmetry breaking and quantum geometric contribution. The photoresponse depends substantially on the polarization state of the excitation light and is highly tunable by external electric fields. This wide tunability in quantum geometric properties enables us to use a convolutional neural network15,16 to achieve full-Stokes polarimetry together with wavelength detection simultaneously, using only one single TDBG device with a subwavelength footprint of merely 3 × 3 µm2. Our work not only reveals the unique role of moiré engineered quantum geometry in tunable nonlinear light-matter interactions but also identifies a pathway for future intelligent sensing technologies in an extremely compact, on-chip manner.

A tissue-like neurotransmitter sensor for the brain and gut.

Neurotransmitters play essential roles in regulating neural circuit dynamics both in the central nervous system as well as at the peripheral, including the gastrointestinal tract1-3. Their real-time monitoring will offer critical information for understanding neural function and diagnosing disease1-3. However, bioelectronic tools to monitor the dynamics of neurotransmitters in vivo, especially in the enteric nervous systems, are underdeveloped. This is mainly owing to the limited availability of biosensing tools that are capable of examining soft, complex and actively moving organs. Here we introduce a tissue-mimicking, stretchable, neurochemical biological interface termed NeuroString, which is prepared by laser patterning of a metal-complexed polyimide into an interconnected graphene/nanoparticle network embedded in an elastomer. NeuroString sensors allow chronic in vivo real-time, multichannel and multiplexed monoamine sensing in the brain of behaving mouse, as well as measuring serotonin dynamics in the gut without undesired stimulations and perturbing peristaltic movements. The described elastic and conformable biosensing interface has broad potential for studying the impact of neurotransmitters on gut microbes, brain-gut communication and may ultimately be extended to biomolecular sensing in other soft organs across the body.

Scalable production of high-performing woven lithium-ion fibre batteries.

Fibre lithium-ion batteries are attractive as flexible power solutions because they can be woven into textiles, offering a convenient way to power future wearable electronics1-4. However, they are difficult to produce in lengths of more than a few centimetres, and longer fibres were thought to have higher internal resistances3,5 that compromised electrochemical performance6,7. Here we show that the internal resistance of such fibres has a hyperbolic cotangent function relationship with fibre length, where it first decreases before levelling off as length increases. Systematic studies confirm that this unexpected result is true for different fibre batteries. We are able to produce metres of high-performing fibre lithium-ion batteries through an optimized scalable industrial process. Our mass-produced fibre batteries have an energy density of 85.69 watt hour per kilogram (typical values8 are less than 1 watt hour per kilogram), based on the total weight of a lithium cobalt oxide/graphite full battery, including packaging. Its capacity retention reaches 90.5% after 500 charge-discharge cycles and 93% at 1C rate (compared with 0.1C rate capacity), which is comparable to commercial batteries such as pouch cells. Over 80 per cent capacity can be maintained after bending the fibre for 100,000 cycles. We show that fibre lithium-ion batteries woven into safe and washable textiles by industrial rapier loom can wirelessly charge a cell phone or power a health management jacket integrated with fibre sensors and a textile display.

The ergodicity question when imaging DNA conformation using liquid cell electron microscopy.

Assessing the ergodicity of graphene liquid cell electron microscope measurements, we report that loop states of circular DNA interconvert reversibly and that loop numbers follow the Boltzmann distribution expected for this molecule in bulk solution, provided that the electron dose is low (80-keV electron energy and electron dose rate 1-20 e- Å-2 s-1). This imaging technique appears to act as a "slow motion" camera that reveals equilibrated distributions by imaging the time average of a few molecules without the need to image a spatial ensemble.

Graphene sandwich-based biological specimen preparation for cryo-EM analysis.

High-quality specimen preparation plays a crucial role in cryo-electron microscopy (cryo-EM) structural analysis. In this study, we have developed a reliable and convenient technique called the graphene sandwich method for preparing cryo-EM specimens. This method involves using two layers of graphene films that enclose macromolecules on both sides, allowing for an appropriate ice thickness for cryo-EM analysis. The graphene sandwich helps to mitigate beam-induced charging effect and reduce particle motion compared to specimens prepared using the traditional method with graphene support on only one side, therefore improving the cryo-EM data quality. These advancements may open new opportunities to expand the use of graphene in the field of biological electron microscopy.

Engineering bionanoreactor in bacteria for efficient hydrogen production.

Hydrogen production through water splitting is a vital strategy for renewable and sustainable clean energy. In this study, we developed an approach integrating nanomaterial engineering and synthetic biology to establish a bionanoreactor system for efficient hydrogen production. The periplasmic space (20 to 30 nm) of an electroactive bacterium, Shewanella oneidensis MR-1, was engineered to serve as a bionanoreactor to enhance the interaction between electrons and protons, catalyzed by hydrogenases for hydrogen generation. To optimize electron transfer, we used the microbially reduced graphene oxide (rGO) to coat the electrode, which improved the electron transfer from the electrode to the cells. Native MtrCAB protein complex on S. oneidensis and self-assembled iron sulfide (FeS) nanoparticles acted in tandem to facilitate electron transfer from an electrode to the periplasm. To enhance proton transport, S. oneidensis MR-1 was engineered to express Gloeobacter rhodopsin (GR) and the light-harvesting antenna canthaxanthin. This led to efficient proton pumping when exposed to light, resulting in a 35.6% increase in the rate of hydrogen production. The overexpression of native [FeFe]-hydrogenase further improved the hydrogen production rate by 56.8%. The bionanoreactor engineered in S. oneidensis MR-1 achieved a hydrogen yield of 80.4 µmol/mg protein/day with a Faraday efficiency of 80% at a potential of -0.75 V. This periplasmic bionanoreactor combines the strengths of both nanomaterial and biological components, providing an efficient approach for microbial electrosynthesis.

Uniform thin ice on ultraflat graphene for high-resolution cryo-EM.

Cryo-electron microscopy (cryo-EM) visualizes the atomic structure of macromolecules that are embedded in vitrified thin ice at their close-to-native state. However, the homogeneity of ice thickness, a key factor to ensure high image quality, is poorly controlled during specimen preparation and has become one of the main challenges for high-resolution cryo-EM. Here we found that the uniformity of thin ice relies on the surface flatness of the supporting film, and developed a method to use ultraflat graphene (UFG) as the support for cryo-EM specimen preparation to achieve better control of vitreous ice thickness. We show that the uniform thin ice on UFG improves the image quality of vitrified specimens. Using such a method we successfully determined the three-dimensional structures of hemoglobin (64 kDa), α-fetoprotein (67 kDa) with no symmetry, and streptavidin (52 kDa) at a resolution of 3.5 Å, 2.6 Å and 2.2 Å, respectively. Furthermore, our results demonstrate the potential of UFG for the fields of cryo-electron tomography and structure-based drug discovery.

Carbon Nanomaterial Fluorescent Probes and Their Biological Applications.

Fluorescent carbon nanomaterials have broadly useful chemical and photophysical attributes that are conducive to applications in biology. In this review, we focus on materials whose photophysics allow for the use of these materials in biomedical and environmental applications, with emphasis on imaging, biosensing, and cargo delivery. The review focuses primarily on graphitic carbon nanomaterials including graphene and its derivatives, carbon nanotubes, as well as carbon dots and carbon nanohoops. Recent advances in and future prospects of these fields are discussed at depth, and where appropriate, references to reviews pertaining to older literature are provided.

Layered nanocomposites by shear-flow-induced alignment of nanosheets.

Biological materials, such as bones, teeth and mollusc shells, are well known for their excellent strength, modulus and toughness1-3. Such properties are attributed to the elaborate layered microstructure of inorganic reinforcing nanofillers, especially two-dimensional nanosheets or nanoplatelets, within a ductile organic matrix4-6. Inspired by these biological structures, several assembly strategies-including layer-by-layer4,7,8, casting9,10, vacuum filtration11-13 and use of magnetic fields14,15-have been used to develop layered nanocomposites. However, how to produce ultrastrong layered nanocomposites in a universal, viable and scalable manner remains an open issue. Here we present a strategy to produce nanocomposites with highly ordered layered structures using shear-flow-induced alignment of two-dimensional nanosheets at an immiscible hydrogel/oil interface. For example, nanocomposites based on nanosheets of graphene oxide and clay exhibit a tensile strength of up to 1,215 ± 80 megapascals and a Young's modulus of 198.8 ± 6.5 gigapascals, which are 9.0 and 2.8 times higher, respectively, than those of natural nacre (mother of pearl). When nanosheets of clay are used, the toughness of the resulting nanocomposite can reach 36.7 ± 3.0 megajoules per cubic metre, which is 20.4 times higher than that of natural nacre; meanwhile, the tensile strength is 1,195 ± 60 megapascals. Quantitative analysis indicates that the well aligned nanosheets form a critical interphase, and this results in the observed mechanical properties. We consider that our strategy, which could be readily extended to align a variety of two-dimensional nanofillers, could be applied to a wide range of structural composites and lead to the development of high-performance composites.

Nanomechanoelectrical approach to highly sensitive and specific label-free DNA detection.

Electronic detection of DNA oligomers offers the promise of rapid, miniaturized DNA analysis across various biotechnological applications. However, known all-electrical methods, which solely rely on measuring electrical signals in transducers during probe-target DNA hybridization, are prone to nonspecific electrostatic and electrochemical interactions, subsequently limiting their specificity and detection limit. Here, we demonstrate a nanomechanoelectrical approach that delivers ultra-robust specificity and a 100-fold improvement in detection limit. We drive nanostructural DNA strands tethered to a graphene transistor to oscillate in an alternating electric field and show that the transistor-current spectra are characteristic and indicative of DNA hybridization. We find that the inherent difference in pliability between unpaired and paired DNA strands leads to the spectral characteristics with minimal influence from nonspecific electrostatic and electrochemical interactions, resulting in high selectivity and sensitivity. Our results highlight the potential of high-performance DNA analysis based on miniaturized all-electronic settings.

In pursuit of degenerative brain disease diagnosis: Dementia biomarkers detected by DNA aptamer-attached portable graphene biosensor.

Dementia is a brain disease which results in irreversible and progressive loss of cognition and motor activity. Despite global efforts, there is no simple and reliable diagnosis or treatment option. Current diagnosis involves indirect testing of commonly inaccessible biofluids and low-resolution brain imaging. We have developed a portable, wireless readout-based Graphene field-effect transistor (GFET) biosensor platform that can detect viruses, proteins, and small molecules with single-molecule sensitivity and specificity. We report the detection of three important amyloids, namely, Amyloid beta (Aß), Tau (τ), and α-Synuclein (αS) using DNA aptamer nanoprobes. These amyloids were isolated, purified, and characterized from the autopsied brain tissues of Alzheimer's Disease (AD) and Parkinson's Disease (PD) patients. The limit of detection (LoD) of the sensor is 10 fM, 1-10 pM, 10-100 fM for Aß, τ, and αS, respectively. Synthetic as well as autopsied brain-derived amyloids showed a statistically significant sensor response with respect to derived thresholds, confirming the ability to define diseased vs. nondiseased states. The detection of each amyloid was specific to their aptamers; Aß, τ, and αS peptides when tested, respectively, with aptamers nonspecific to them showed statistically insignificant cross-reactivity. Thus, the aptamer-based GFET biosensor has high sensitivity and precision across a range of epidemiologically significant AD and PD variants. This portable diagnostic system would allow at-home and POC testing for neurodegenerative diseases globally.

Edge states in rationally terminated honeycomb structures.

Consider the tight binding model of graphene, sharply terminated along an edge l parallel to a direction of translational symmetry of the underlying period lattice. We classify such edges l into those of "zigzag type" and those of "armchair type," generalizing the classical zigzag and armchair edges. We prove that zero-energy/flat-band edge states arise for edges of zigzag type, but never for those of armchair type. We exhibit explicit formulae for flat-band edge states when they exist. We produce strong evidence for the existence of dispersive (nonflat) edge state curves of nonzero energy for most l.

Anomalous water transport in narrow-diameter carbon nanotubes.

Carbon nanotubes (CNTs) mimicking the structure of aquaporins support fast water transport, making them strong candidates for building next-generation high-performance membranes for water treatment. The diffusion and transport behavior of water through CNTs or nanoporous graphene can be fundamentally different from those of bulk water through a macroscopic tube. To date, the nanotube-length-dependent physical transport behavior of water is still largely unexplored. Herein, on the basis of molecular dynamics simulations, we show that the flow rate of water through 0.83-nm-diameter (6,6) and 0.96-nm-diameter (7,7) CNTs exhibits anomalous transport behavior, whereby the flow rate increases markedly first and then either slowly decreases or changes slightly as the CNT length l increases. The critical range of l for the flow-rate transition is 0.37 to 0.5 nm. This anomalous water transport behavior is attributed to the l-dependent mechanical stability of the transient hydrogen-bonding chain that connects water molecules inside and outside the CNTs and bypasses the CNT orifice. The results unveil a microscopic mechanism governing water transport through subnanometer tubes, which has important implications for nanofluidic manipulation.

Precise radiocarbon determination in radioactive waste by a laser-based spectroscopic technique.

The precise and accurate determination of the radionuclide inventory in radioactive waste streams, including those generated during nuclear decommissioning, is a key aspect in establishing the best-suited nuclear waste management and disposal options. Radiocarbon ([Formula: see text]) is playing a crucial role in this scenario because it is one of the so-called difficult to measure isotopes; currently, [Formula: see text] analysis requires complex systems, such as accelerator mass spectrometry (AMS) or liquid scintillation counting (LSC). AMS has an outstanding limit of detection, but only a few facilities are available worldwide; LSC, which can have similar performance, is more widespread, but sample preparation can be nontrivial. In this paper, we demonstrate that the laser-based saturated-absorption cavity ring-down (SCAR) spectroscopic technique has several distinct advantages and represents a mature and accurate alternative for [Formula: see text] content determination in nuclear waste. As a proof-of-principle experiment, we show consistent results of AMS and SCAR for samples of concrete and graphite originating from nuclear installations. In particular, we determined mole fractions of 1.312(9) F[Formula: see text] and 30.951(7) F[Formula: see text] corresponding to ∼1.5 and 36.2 parts per trillion (ppt), respectively, for two different graphite samples originating from different regions of the Adiabatic Resonance Crossing activator prototype installed on one irradiation line of an MC40 Scanditronix cyclotron. Moreover, we measure a mole fraction of 0.593(8) F[Formula: see text] ([Formula: see text] ppt) from a concrete sample originating from an external wall of the Ispra-1 nuclear research reactor currently in the decommissioning phase.

Anode electrolysis of sulfides.

Traditional sulfide metallurgy produces harmful sulfur dioxide and is energy intensive. To this end, we develop an anode electrolysis approach in molten salt by which sulfide is electrochemically split into sulfur gas at a graphite inert anode while releasing metal ions that diffuse toward and are deposited at the cathode. The anodic splitting dictates the "sulfide-to-metal ion and sulfur gas" conversion that makes the reaction recur continuously. Using this approach, Cu2S is converted to sulfur gas and Cu in molten LiCl-KCl at 500 °C with a current efficiency of 99% and energy consumption of 0.420 kWh/kg-Cu (only considering the electricity for electrolysis). Besides Cu2S, the anode electrolysis can extract Cu from Cu matte that is an intermediate product from the traditional sulfide smelting process. More broadly, Fe, Ni, Pb, and Sb are extracted from FeS, CuFeS2, NiS, PbS, and Sb2S3, providing a general electrochemical method for sulfide metallurgy.

On the mechanisms of ion adsorption to aqueous interfaces: air-water vs. oil-water.

The adsorption of ions to water-hydrophobe interfaces influences a wide range of phenomena, including chemical reaction rates, ion transport across biological membranes, and electrochemical and many catalytic processes; hence, developing a detailed understanding of the behavior of ions at water-hydrophobe interfaces is of central interest. Here, we characterize the adsorption of the chaotropic thiocyanate anion (SCN-) to two prototypical liquid hydrophobic surfaces, water-toluene and water-decane, by surface-sensitive nonlinear spectroscopy and compare the results against our previous studies of SCN- adsorption to the air-water interface. For these systems, we observe no spectral shift in the charge transfer to solvent spectrum of SCN-, and the Gibb's free energies of adsorption for these three different interfaces all agree within error. We employed molecular dynamics simulations to develop a molecular-level understanding of the adsorption mechanism and found that the adsorption for SCN- to both water-toluene and water-decane interfaces is driven by an increase in entropy, with very little enthalpic contribution. This is a qualitatively different mechanism than reported for SCN- adsorption to the air-water and graphene-water interfaces, wherein a favorable enthalpy change was the main driving force, against an unfavorable entropy change.

Muscle degeneration in chronic massive rotator cuff tears of the shoulder: Addressing the real problem using a graphene matrix.

Massive rotator cuff tears (MRCTs) of the shoulder cause disability and pain among the adult population. In chronic injuries, the tendon retraction and subsequently the loss of mechanical load lead to muscle atrophy, fat accumulation, and fibrosis formation over time. The intrinsic repair mechanism of muscle and the successful repair of the torn tendon cannot reverse the muscle degeneration following MRCTs. To address these limitations, we developed an electroconductive matrix by incorporating graphene nanoplatelets (GnPs) into aligned poly(l-lactic acid) (PLLA) nanofibers. This study aimed to understand 1) the effects of GnP matrices on muscle regeneration and inhibition of fat formation in vitro and 2) the ability of GnP matrices to reverse muscle degenerative changes in vivo following an MRCT. The GnP matrix significantly increased myotube formation, which can be attributed to enhanced intracellular calcium ions in myoblasts. Moreover, the GnP matrix suppressed adipogenesis in adipose-derived stem cells. These results supported the clinical effects of the GnP matrix on reducing fat accumulation and muscle atrophy. The histological evaluation showed the potential of the GnP matrix to reverse muscle atrophy, fat accumulation, and fibrosis in both supraspinatus and infraspinatus muscles at 24 and 32 wk after the chronic MRCTs of the rat shoulder. The pathological evaluation of internal organs confirmed the long-term biocompatibility of the GnP matrix. We found that reversing muscle degenerative changes improved the morphology and tensile properties of the tendon compared with current surgical techniques. The long-term biocompatibility and the ability of the GnP matrix to treat muscle degeneration are promising for the realization of MRCT healing and regeneration.