Single-molecular protein-based bioelectronics via electronic transport: fundamentals, devices and applications.

Biomolecular electronics is a rapidly growing multidisciplinary field that combines biology, nanoscience, and engineering to bridge the two important fields of life sciences and molecular electronics. Proteins are remarkable for their ability to recognize molecules and transport electrons, making the integration of proteins into electronic devices a long sought-after goal and leading to the emergence of the field of protein-based bioelectronics, also known as proteotronics. This field seeks to design and create new biomolecular electronic platforms that allow for the understanding and manipulation of protein-mediated electronic charge transport and related functional applications. In recent decades, there have been numerous reports on protein-based bioelectronics using a variety of nano-gapped electrical devices and techniques at the single molecular level, which are not achievable with conventional ensemble approaches. This review focuses on recent advances in physical electron transport mechanisms, device fabrication methodologies, and various applications in protein-based bioelectronics. We discuss the most recent progress of the single or few protein-bridged electrical junction fabrication strategies, summarise the work on fundamental and functional applications of protein bioelectronics that enable high and dynamic electron transport, and highlight future perspectives and challenges that still need to be addressed. We believe that this specific review will stimulate the interdisciplinary research of topics related to protein-related bioelectronics, and open up new possibilities for single-molecule biophysics and biomedicine.

Single-Molecule Surface-Enhanced Raman Spectroscopy.

Single-molecule surface-enhanced Raman spectroscopy (SM-SERS) has the potential to detect single molecules in a non-invasive, label-free manner with high-throughput. SM-SERS can detect chemical information of single molecules without statistical averaging and has wide application in chemical analysis, nanoelectronics, biochemical sensing, etc. Recently, a series of unprecedented advances have been realized in science and application by SM-SERS, which has attracted the interest of various fields. In this review, we first elucidate the key concepts of SM-SERS, including enhancement factor (EF), spectral fluctuation, and experimental evidence of single-molecule events. Next, we systematically discuss advanced implementations of SM-SERS, including substrates with ultra-high EF and reproducibility, strategies to improve the probability of molecules being localized in hotspots, and nonmetallic and hybrid substrates. Then, several examples for the application of SM-SERS are proposed, including catalysis, nanoelectronics, and sensing. Finally, we summarize the challenges and future of SM-SERS. We hope this literature review will inspire the interest of researchers in more fields.

Instrument response effects on the retrieval of oceanic lidar.

Seawater properties can be retrieved from oceanic lidar returns. However, the actual returns include the ideal returns convolved by the instrument response, which inevitably introduces retrieval error. In this paper, instrument response effects on the retrieval of oceanic lidar are analyzed from different aspects. The results demonstrate that the retrieval of the lidar attenuation coefficient near the water surface is affected by the instrument response in homogeneous water. Considering the ratio of the signal distortion region (relative error of attenuation >10%) to the maximum detection depth (three dynamic ranges) is less than 20%, the pulse width of the instrument response should be less than 10-0.042(Kd)-2+0.709(Kd)-1+1.136ns. In addition, an average relative error of 55% will be introduced to the retrieval of phytoplankton layer thickness in the stratified water, which can be reduced to 6% after correcting for the influence of the instrument response. However, a relative error greater than 20% still exists when the instrument response length is two times larger than the layer thickness. These conclusions provide guidelines to a future design of oceanic lidar.

miR-30a can inhibit DNA replication by targeting RPA1 thus slowing cancer cell proliferation.

Cell proliferation was inhibited following forced over-expression of miR-30a in the ovary cancer cell line A2780DX5 and the gastric cancer cell line SGC7901R. Interestingly, miR-30a targets the DNA replication protein RPA1, hinders the replication of DNA and induces DNA fragmentation. Furthermore, ataxia telangiectasia mutated (ATM) and checkpoint kinase 2 (CHK2) were phosphorylated after DNA damage, which induced p53 expression, thus triggering the S-phase checkpoint, arresting cell cycle progression and ultimately initiating cancer cell apoptosis. Therefore, forced miR-30a over-expression in cancer cells can be a potential way to inhibit tumour development.

Flexible and Transparent Surface-Enhanced Raman Scattering (SERS)-Active Metafilm for Visualizing Trace Molecules via Raman Spectral Mapping.

Raman spectral mapping is a powerful tool for directly visualizing the composition, structure, and distribution of molecules on any surface of interest. However, one major limitation of Raman mapping is its overlong imaging time caused by the intrinsic weak Raman signal. Here, we developed a fast Raman imaging approach based on a flexible and transparent surface-enhanced Raman scattering (SERS)-active metafilm. This particular SERS substrate can be conformably attached to a sample surface to enhance the Raman signal of analytes and the good optical transparency allow excitation and collection of signal from the backside of the substrate. Therefore, by simply attaching it to the surface of interest, a fast Raman imaging can be realized. We noticed that the imaging speed can be increased by several orders of magnitude, compared to a conventional Raman mapping approach. Importantly, the proposed approach required little or no sample preparation and exhibited good generalizability that can be performed perfectly on different surfaces. It is believed that the proposed methodology will provide new trends for chemical imaging using Raman microscopy.

The graphene/nucleic acid nanobiointerface.

The combination of nanomaterials with biomolecules yields functional nanostructured biointerfaces with synergistic properties and functions. Owing to a unique combination of its crystallographic and electronic structure, graphene and its derivatives exhibit several superior and typical properties, and has emerged as an attractive candidate for the fabrication of novel nanobiointerfaces with different kinds of unique applications. As is known, nucleic acids are stable and can easily handle modification, and can recognize a wide range of targets with high selectivity, specificity, and affinity. The integration of nucleic acids with graphene-based materials has been substantially advanced over the past few years, achieving amazing properties and functions, thereby exhibiting attractive potential applications in biosensing, diagnostics, drug screening and biomedicine. Herein, this review addresses the recent progress on the design and fabrication of graphene/nucleic acid nanostructured biointerfaces, and the fundamental understanding of their interfacial properties, as well as the various nanobiotechnological applications. To begin with, we summarize the basic features of the graphene and nucleic acid-based nanobiointerface, especially the interfacial interaction mechanism and the resulting biological effects. Then, the fabrication and characterization methodology of graphene and nucleic acid-based nanobiointerfaces are discussed. Next, particular emphasis is directed towards the exploration of their biosensing and biomedical applications, including small molecule detection, protein and DNA sensing/sequencing, as well as gene delivery and therapy. Finally, some significant prospects, further opportunities and challenges in this emerging field are also suggested.

A Meta-Analysis of the Efficacy and Safety of Using Oil Massage to Promote Infant Growth.

UNLABELLED: The synthesizing evidence on the effectiveness of using oil massage to promote the growth of infants is still lacking. This paper aims to determine whether oil massage can promote the physical and neurobehavioral growth of infants according to variables and to evaluate whether oil massage is safe for infant skin. ELIGIBILITY CRITERIA: The randomized controlled trials, clinical controlled trials and quasi-experimentally designed trials published prior to or in 2014 were searched according to predetermined inclusion criteria and exclusion criteria in Medline, PubMed, Ovid, the Cochran Library, and Chinese databases, including the China National Knowledge Infrastructure, Wan Fang database and VIP journal integration platform. Besides, the grey lectures were searched as well through Open Grey, GrayLIT Network and Clinical Trials.gov. SAMPLE: Eight studies out of 625 retrieved articles were eligible for inclusion. RESULTS: Oil massage increased the infant weights, lengths and head circumferences. However, it did not promote a significant advantage in neurobehavioral scores or cause a significant risk of adverse skin reactions. IMPLICATIONS: The core mechanisms and standard procedures of oil massage as well as the preferred oil type should be the focus of future nursing practice and research. CONCLUSIONS: Oil massage may effectively improve the physical growth of infants, and it presents a limited risk of adverse skin reactions. However, the relationship between neurodevelopment and oil massage requires further study.

Nitrite-triggered surface plasmon-assisted catalytic conversion of p-aminothiophenol to p,p'-dimercaptoazobenzene on gold nanoparticle: surface-enhanced Raman scattering investigation and potential for nitrite detection.

The stunning large enhancement factor (∼10(8)) of the surface-enhanced Raman scattering (SERS) effect leads people to wonder about the underlying enhancement mechanisms of the effect. But, a strong evidence of the existence of one commonly accepted mechanism (chemical enhancement), the origin of the symbolic "b2" bands (ca. 1140,1390, 1432 cm(-1)) of p-aminothiophenol (PATP), was recently found to be a false explanation, which were actually arisen from the product of a surface plasmon-assisted coupling reaction of PATP, p,p'-dimercaptoazobenzene (DMAB). However, the debate is far from over, especially because the mechanism of the above reaction has not been fully understood yet. In this paper, we for the first time report a new surface plasmon-assisted catalytic conversion of PATP to DMAB that NO2(-) ions can trigger the formation of DMAB on gold nanoparticles (GNPs) suspension under light illumination. The mechanism of the conversion is also discussed. All relevant data suggest the nitrite-triggered conversion of PATP to DMAB on GNPs is a surface plasmon-assisted oxidation reaction, involving transfer of multiple electrons from PATP to NO2(-) (electron acceptors) and protons, leading to the formation of DMAB. The proposed mechanisms may also help to understand the unclear surface plasmon-assisted catalytic coupling of PATP on the SERS substrates. Furthermore, inspired by the high selectivity of the above nitrite-triggered catalysis reaction, a simple and fast nitrite screening method was also developed, exhibiting good sensitivity. Considering other advantages of the assay, such as rapidness, simplicity of the detection procedures, and requirement of no sample pretreatment, it is a promising method for on-site fast screening or point-of-care application.

Ultrasensitive Detection of Testosterone Using Microring Resonator with Molecularly Imprinted Polymers.

We report ultrasensitive and highly selective detection of testosterone based on microring resonance sensor using molecularly imprinted polymers (MIP). A silicon-on-insulator (SOI) micoring resonator was modified by MIP films (MIPs) on a surface. The MIPs was synthesized by thermopolymerization using methacrylic acid as functional monomer and ethylene glycol dimethacrylate as crosslinking agent. The concentration of detected testosterone varies from 0.05 ng/mL to 10 ng/mL. The detection limit reaches 48.7 pg/mL. Ultrahigh sensitivity, good specificity and reproducibility have been demonstrated, indicating the great potential of making a cost effective and easy to operate lab-on-Chip and down scaling micro-fluidics devices in biosensing.

DNA-Encoded Tuning of Geometric and Plasmonic Properties of Nanoparticles Growing from Gold Nanorod Seeds.

Systematically controlling the morphology of nanoparticles, especially those growing from gold nanorod (AuNR) seeds, are underexplored; however, the AuNR and its related morphologies have shown promises in many applications. Herein we report the use of programmable DNA sequences to control AuNR overgrowth, resulting in gold nanoparticles varying from nanodumbbell to nanooctahedron, as well as shapes in between, with high yield and reproducibility. Kinetic studies revealed two representative pathways for the shape control evolving into distinct nanostructures. Furthermore, the geometric and plasmonic properties of the gold nanoparticles could be precisely controlled by adjusting the base compositions of DNA sequences or by introducing phosphorothioate modifications in the DNA. As a result, the surface plasmon resonance (SPR) peaks of the nanoparticles can be fine-tuned in a wide range, from visible to second near-infrared (NIR-II) region beyond 1000 nm.

High-sensitivity optical biosensor based on cascaded Mach-Zehnder interferometer and ring resonator using Vernier effect.

We demonstrate an ultrahigh sensitivity silicon photonic biosensor based on cascaded Mach-Zehnder interferometer (MZI) and ring resonator with the Vernier effect using wavelength interrogation. Experimental results show that the sensitivities reached 2870 nm/RIU and 21,500 nm/RIU for MZI sensor and MZI-ring sensor, respectively. A biosensing application was demonstrated by monitoring the interaction between goat and antigoat immunoglobulin G (IgG) pairs. The measured results show that 1 ng/ml IgG resulted in 0.035 nm and 0.5 nm wavelength shift for MZI sensor and MZI-ring sensor, respectively. This high performance sensor is promising for medical diagnostic applications.

Toehold-initiated rolling circle amplification for visualizing individual microRNAs in situ in single cells.

The ability to quantitate and visualize microRNAs (miRNAs) in situ in single cells would greatly facilitate the elucidation of miRNA-mediated regulatory circuits and their disease associations. A toehold-initiated strand-displacement process was used to initiate rolling circle amplification of specific miRNAs, an approach that achieves both stringent recognition and in situ amplification of the target miRNA. This assay, termed toehold-initiated rolling circle amplification (TIRCA), can be utilized to identify miRNAs at physiological temperature with high specificity and to visualize individual miRNAs in situ in single cells within 3 h. TIRCA is a competitive candidate technique for in situ miRNA imaging and may help us to understand the role of miRNAs in cellular processes and human diseases in more detail.

The J bs-5YP peptide can alleviate dementia in senile mice by restoring the transcription of Slc40a1 to secrete the excessive iron from brain.

INTRODUCTION: With age and ATP decrease in the body, the transcription factors hypophosphorylation weakens the transcription of Slc40a1 and hinders the expression of the iron discharger ferroportin. This may lead to iron accumulation in the brain and the catalysis of free radicals that damage cerebral neurons and eventually lead to Alzheimer's disease (AD). OBJECTIVES: To prevent AD caused by brain iron excretion disorders and reveal the mechanism of J bs-5YP peptide restoring ferroportin. METHODS: We prepared J bs-YP peptide and administered it to the senile mice with dementia. Then, the intelligence of the mice was tested using a Morris Water Maze. The ATP content in the body was detected using the ATP hydrophysis and Phosphate precipitation method. The activation of Slc40a1 transcription was assayed with ATAC seq and the ferroportin, as well as the phosphorylation levels of Ets1 in brain were detected by Western Blot. RESULTS: The phosphorylation level of Ets1in brain was enhanced, and subsequently, the transcription of Slc40a1 was activated and ferroportin was increased in the brain, the levels of iron and free radicals were reduced, with the neurons protection, and the dementia was ultimately alleviated in the senile mice. CONCLUSION: J bs-5YP can recover the expression of ferroportin to excrete excessive iron in the brain of senile mice with dementia by enhancing the transcription of Slc40a1 via phosphorylating Ets1, revealing the potential of J bs-5YP as a drug to alleviate senile dementia.

DNA detection using plasmonic enhanced near-infrared photoluminescence of gallium arsenide.

Efficient near-infrared detection of specific DNA with single nucleotide polymorphism selectivity is important for diagnostics and biomedical research. Herein, we report the use of gallium arsenide (GaAs) as a sensing platform for probing DNA immobilization and targeting DNA hybridization, resulting in ∼8-fold enhanced GaAs photoluminescence (PL) at ∼875 nm. The new signal amplification strategy, further coupled with the plasmonic effect of Au nanoparticles, is capable of detecting DNA molecules with a detection limit of 0.8 pM and selectivity against single base mismatches. Such an ultrasensitive near-infrared sensor can find a wide range of biochemical and biomedical applications.

Fabrication of electron tunneling probes for measuring single-protein conductance.

Studying the electrical properties of individual proteins is a prominent research area in the field of bioelectronics. Electron tunnelling or quantum mechanical tunnelling (QMT) probes can act as powerful tools for investigating the electrical properties of proteins. However, current fabrication methods for these probes often have limited reproducibility, unreliable contact or inadequate binding of proteins onto the electrodes, so better solutions are required. Here, we detail a generalizable and straightforward set of instructions for fabricating simple, nanopipette-based, tunnelling probes, suitable for measuring conductance in single proteins. Our QMT probe is based on a high-aspect-ratio dual-channel nanopipette that integrates a pair of gold tunneling electrodes with a gap of less than 5 nm, fabricated via the pyrolytic deposition of carbon followed by the electrochemical deposition of gold. The gold tunneling electrodes can be functionalized using an extensive library of available surface modifications to achieve single-protein-electrode contact. We use a biotinylated thiol modification, in which a biotin-streptavidin-biotin bridge is used to form the single-protein junction. The resulting protein-coupled QMT probes enable the stable electrical measurement of the same single protein in solution for up to several hours. We also describe the analysis method used to interpret time-dependent single-protein conductance measurements, which can provide essential information for understanding electron transport and exploring protein dynamics. The total time required to complete the protocol is ~33 h and it can be carried out by users trained in less than 24 h.

Multiplexed detection of viral antigen and RNA using nanopore sensing and encoded molecular probes.

We report on single-molecule nanopore sensing combined with position-encoded DNA molecular probes, with chemistry tuned to simultaneously identify various antigen proteins and multiple RNA gene fragments of SARS-CoV-2 with high sensitivity and selectivity. We show that this sensing strategy can directly detect spike (S) and nucleocapsid (N) proteins in unprocessed human saliva. Moreover, our approach enables the identification of RNA fragments from patient samples using nasal/throat swabs, enabling the identification of critical mutations such as D614G, G446S, or Y144del among viral variants. In particular, it can detect and discriminate between SARS-CoV-2 lineages of wild-type B.1.1.7 (Alpha), B.1.617.2 (Delta), and B.1.1.539 (Omicron) within a single measurement without the need for nucleic acid sequencing. The sensing strategy of the molecular probes is easily adaptable to other viral targets and diseases and can be expanded depending on the application required.

Colorimetric and ultrasensitive bioassay based on a dual-amplification system using aptamer and DNAzyme.

Rapid detection of ultralow amount of biomarkers in a biologically complex mixture remains a major challenge. Herein, we report a novel aptamer-based protein detection assay that integrates two signal amplification processes, namely, polymerase-mediated rolling-circle amplification (RCA) and DNA enzyme-catalyzed colorimetric reaction. The target biomarker is captured in a sandwich assay by primary aptamer-functionalized microbeads (MBs) and a secondary aptamer that is connected to a RCA primer/circular template complex. RCA reaction, which amplifies the single biomarker binding events by a factor of hundreds to thousands (the first amplification) produces a long DNA molecule containing multiple DNAzyme units. The peroxidase-like DNAzyme catalyzes the oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (the second amplification), which generates a blue-green colorimetric signal. This new biosensing platform permits the ultrasensitive, label-free, colorimetric detection of biomarker in real time. Using platelet-derived growth factor B-chain (PDGF-BB) as a model system, we demonstrated that our assay can detect a protein marker specifically in a serum-containing medium, at a concentration as low as 0.2 pg/mL in ∼2 h, which rivals traditional assays such as ELISA. We anticipate this simple methodology for biomarker detection can find utility in point-of-care applications.

Discovery of the DNA "genetic code" for abiological gold nanoparticle morphologies.

DNA is in control: Different combinations of DNA nucleotides can control the shape and surface roughness of gold nanoparticles during their synthesis. These nanoparticles were synthesized in the presence of either homogenous oligonucleotides or mixed-base oligonucleotides using gold nanoprisms as seeds. The effect of the individual DNA bases and their combinations on shape control are shown in the figure.

Sub-5 nm nanogap electrodes towards single-molecular biosensing.

Nanogap electrodes (NGEs) with sub-5 nm gap has been widely used in single-molecule sensing and sequencing, with the characteristics of label-free, high sensitivity, rapid detection and low-cost. However, the fabrication of sub-5 nm gap electrodes with high controllability and reproducibility still remains a great challenge that impedes the experimental research and the commercialization of the nanogap device. Here, we review the common currently used fabrication methods of nanogap electrodes, such as gap narrowing deposition, mechanical controllable break junctions and the fabrication methods combined with nanopore or nanochannel. We then highlight the typical applications of nanogap electrodes in biological/chemical sensing fields, including single molecule recognition, single molecule sequencing and chemical kinetics analysis. Finally, the challenges of nanogap electrodes in single molecule sensing/sequencing are outlined and the future directions for sensing perspectives are suggested.

Measuring conductance switching in single proteins using quantum tunneling.

Interpreting the electrical signatures of single proteins in electronic junctions has facilitated a better understanding of the intrinsic properties of proteins that are fundamental to chemical and biological processes. Often, this information is not accessible using ensemble and even single-molecule approaches. In addition, the fabrication of nanoscale single-protein junctions remains challenging as they often require sophisticated methods. We report on the fabrication of tunneling probes, direct measurement, and active control (switching) of single-protein conductance with an external field in solution. The probes allowed us to bridge a single streptavidin molecule to two independently addressable, biotin-terminated electrodes and measure single-protein tunneling response over long periods. We show that charge transport through the protein has multiple conductive pathways that depend on the magnitude of the applied bias. These findings open the door for the reliable fabrication of protein-based junctions and can enable their use in future protein-embedded bioelectronics applications.