Ratoon Stunting Disease of Sugarcane: A Review Emphasizing Detection Strategies and Challenges.

Sugarcane (Saccharum hybrid) is an important cash crop grown in tropical and subtropical countries. Ratoon stunting disease (RSD), caused by a xylem-inhabiting bacterium, Leifsonia xyli subsp. xyli (Lxx) is one of the most economically significant diseases globally. RSD results in severe yield losses because its highly contagious nature and lack of visually identifiable symptoms make it harder to devise an effective management strategy. The efficacy of current management practices is hindered by implementation difficulties caused by lack of resources, high cost, and difficulties in monitoring. Rapid detection of the causal pathogen in vegetative planting material is crucial for sugarcane growers to manage this disease. Several microscopic, serological, and molecular-based methods have been developed and used for detecting the RSD pathogen. Although these methods have been used across the sugarcane industry worldwide to diagnose Lxx, some lack reliability or specificity, are expensive and time-consuming to apply, and most of all, are not suitable for on-farm diagnosis. In recent decades, there has been significant progress in the development of integrated isothermal amplification-based microdevices for accurate human and plant pathogen detection. There is a significant opportunity to develop a novel diagnostic method that integrates nanobiosensing with isothermal amplification within a microdevice format for accurate Lxx detection. In this review, we summarize (i) the historical background and current knowledge of sugarcane ratoon stunting disease, including some aspects related to transmission, pathosystem, and management practices; and (ii) the drawbacks of current diagnostic methods and the potential for application of advanced diagnostics to improve disease management.

Electrocatalytic Microdevice Array Based on Wafer-Scale Conductive Metal-Organic Framework Thin Film for Massive Hydrogen Production.

The synthesis of large-scale 2D conductive metal-organic framework films with tunable thickness is highly desirable but challenging. In this study, an Interface Confinement Self-Assembly Pulling (ICSP) method for in situ synthesis of 4-in. Ni-BHT film on the substrate surface is developed. By modulating the thickness of the confined space, the thickness of Ni-BHT films could be easily varied from 4 to 42 nm. To eliminate interference factors and evaluate the effect of film thickness on the catalytic performance of HER, an electrocatalytic microdevice based on the Ni-BHT film is designed. The effective catalytic thickness of the Ni-BHT film is found to be around 32 nm. Finally, to prepare the electrocatalytic microdevice array, over 100 000 microdevices on a 4-in. Ni-BHT film are integrated. The results show that the microdevice array has good stability and a high hydrogen production rate and could be used to produce large amounts of hydrogen. The wafer-scale 2D conductive metal-organic framework's fabrication greatly advances the practical application of microdevices for massive hydrogen production.

A low-cost and hand-hold PCR microdevice based on water-cooling technology.

Polymerase chain reaction (PCR) has become a powerful tool for detecting various diseases due to its high sensitivity and specificity. However, the long thermocycling time and the bulky system have limited the application of PCR devices in Point-of-care testing. Herein, we have proposed an efficient, low-cost, and hand-hold PCR microdevice, mainly including a control module based on water-cooling technology and an amplification module fabricated by 3D printing. The whole device is tiny and can be easily hand-held with a size of about 110 mm × 100 mm × 40 mm and a weight of about 300 g at a low cost of about $170.83. Based on the water-cooling technology, the device can efficiently perform 30 thermal cycles within 46 min at a heating/cooling rate of 4.0/8.1 ℃/s. To test our instrument, plasmid DNA dilutions were amplified with this device; the results demonstrate successful nucleic acid amplification of the plasmid DNA and exhibit the promise of this device for Point-of-care testing.

Electrochemistry in sensing of molecular interactions of proteins and their behavior in an electric field.

Electrochemical methods can be used not only for the sensitive analysis of proteins but also for deeper research into their structure, transport functions (transfer of electrons and protons), and sensing their interactions with soft and solid surfaces. Last but not least, electrochemical tools are useful for investigating the effect of an electric field on protein structure, the direct application of electrochemical methods for controlling protein function, or the micromanipulation of supramolecular protein structures. There are many experimental arrangements (modalities), from the classic configuration that works with an electrochemical cell to miniaturized electrochemical sensors and microchip platforms. The support of computational chemistry methods which appropriately complement the interpretation framework of experimental results is also important. This text describes recent directions in electrochemical methods for the determination of proteins and briefly summarizes available methodologies for the selective labeling of proteins using redox-active probes. Attention is also paid to the theoretical aspects of electron transport and the effect of an external electric field on the structure of selected proteins. Instead of providing a comprehensive overview, we aim to highlight areas of interest that have not been summarized recently, but, at the same time, represent current trends in the field.

Skill-Learning Method of Dual Peg-in-Hole Compliance Assembly for Micro-Device.

For the dual peg-in-hole compliance assembly task of upper and lower double-hole structural micro-devices, a skill-learning method is proposed. This method combines offline training in a simulation space and online training in a realistic space. In this paper, a dual peg-in-hole model is built according to the results of a force analysis, and contact-point searching methods are provided for calculating the contact force. Then, a skill-learning framework is built based on deep reinforcement learning. Both expert action and incremental action are used in training, and a reward system considers both efficiency and safety; additionally, a dynamic exploration method is provided to improve the training efficiency. In addition, based on experimental data, an online training method is used to optimize the skill-learning model continuously so that the error caused by the deviation in the offline training data from reality can be reduced. The final experiments demonstrate that the method can effectively reduce the contact force while assembling, improve the efficiency and reduce the impact of the change in position and orientation.

Micro-Vision Based High-Precision Space Assembly Approach for Trans-Scale Micro-Device: The CFTA Example.

For assembly of trans-scale micro-device capsule fill tube assemblies (CFTA) for inertial confinement fusion (ICF) targets, a high-precision space assembly approach based on micro-vision is proposed in this paper. The approach consists of three modules: (i) a posture alignment module based on a multi-vision monitoring model that is designed to align two trans-scale micro-parts in 5DOF while one micro-part is in ten microns and the other one is in hundreds of microns; (ii) an insertion depth control module based on a proposed local deformation detection method to control micro-part insertion depth; (iii) a glue mass control module based on simulation research that is designed to control glue mass quantitatively and to bond micro-parts together. A series of experiments were conducted and experimental results reveal that attitude alignment control error is less than ±0.3°, position alignment control error is less than ±5 µm, and insertion depth control error is less than ±5 µm. Deviation of glue spot diameter is controlled at less than 15 µm. A CFTA was assembled based on the proposed approach, the position error in 3D space measured by computerized tomography (CT) is less than 5 µm, and glue spot diameter at the joint is 56 µm. Through volume calculation by the cone calculation formula, the glue mass is about 23 PL when the cone height is half the diameter.

On-Chip Microdevice Unveils Reactant Enrichment Effect Dominated Electrocatalysis Activity in Molecular-Linked Catalysts.

Molecular functionalization has been intensely studied and artificially constructed to advance various electrocatalytic processes. While there is a widely approved charge-doping effect, the underlying action for reactant distribution/transport remains long neglected. Here an on-chip microdevice unravels that the proton enrichment effect at prototypical methylene blue (MB)/MoS2 interfaces rather than charge doping contributes to the hydrogen evolution reaction (HER) activity. Back-gated electrical/electrochemical tests detect quantitatively a strong charge injection from MB to MoS2 realized over diploid carrier density, but these excess carriers are unqualified for the actual enhanced HER activity (from 32 to 125 mA cm-2 at -0.29 V). On-chip electrochemical impedance further certifies that the proton enrichment in the vicinity of MoS2, which is generated by the nucleophilic group of MB, actually dominates the HER activity. This finding uncovers the leading function of molecular-linked catalysts.

Hybrid 3D printed integrated microdevice for the determination of copper ions in human body fluids.

On-site screening of copper ions in body fluid plays a critical role in monitoring human health, especially in heavy pollution areas. In this study, we have developed a hybrid 3D printed integrated microdevice for the determination of copper ions in human body fluids. A fixed and low volume of sample was detected by using the integrated microdevice without any preprocessing. The hybrid channel enables sample uniform mixing and quantitative dilution with buffer solution by inducing the "horseshoe vortex" phenomenon. The electrolytic microcell based on the flow detection system shows a more effective copper ion reaction ratio and, as a result, a better sensitivity. The simulation of the finite element method (FEM) determined the relevant optimum parameters of the hybrid channel and the microcell. The design, fabrication, and detection procedure of the integrated microdevice are here illustrated. The microdevice presented superior detection properties towards copper ions. The calibration curves covered two linear ranges varying from 20 to 100 ppb and 100 to 400 ppb, respectively. The limit of detection was estimated to be 15 ppb (S/N = 3). The relative standard deviation of the peak current measurements was 2.26%. The designed microdevice was further applied to detect copper ions in practical samples (calf serum sample and synthetic human urine sample) using a standard addition method, and the average recovery was found to be 95-104%. The performance of copper ion detection with the integrated microdevice was consistent with that of the inductively coupled plasma mass spectrometry (ICP-MS) in the same practical samples, demonstrating significant practicality in the test of body fluidics. The portable integrated microdevice is an excellent choice for on-site detection and has a promising prospect in the point-of-care testing (POCT) applications.

Recent advances in techniques for fabrication and characterization of nanogap biosensors: A review.

Nanogap biosensors have fascinated researchers due to their excellent electrical properties. Nanogap biosensors comprise three arrays of electrodes that form nanometer-size gaps. The sensing gaps have become the major building blocks of several sensing applications, including bio- and chemosensors. One of the advantages of nanogap biosensors is that they can be fabricated in nanoscale size for various downstream applications. Several studies have been conducted on nanogap biosensors, and nanogap biosensors exhibit potential material properties. The possibilities of combining these unique properties with a nanoscale-gapped device and electrical detection systems allow excellent and potential prospects in biomolecular detection. However, their fabrication is challenging as the gap is becoming smaller. It includes high-cost, low-yield, and surface phenomena to move a step closer to the routine fabrications. This review summarizes different feasible techniques in the fabrication of nanogap electrodes, such as preparation by self-assembly with both conventional and nonconventional approaches. This review also presents a comprehensive analysis of the fabrication, potential applications, history, and the current status of nanogap biosensors with a special focus on nanogap-mediated bio- and chemical sonsors.

Dual-Regulation of Defect Sites and Vertical Conduction by Spiral Domain for Electrocatalytic Hydrogen Evolution.

Insufficient active sites and weak vertical conduction are the intrinsic factors that restrict the electrocatalytic HER for transition-metal dichalcogenides. As a prototype, we proposed a model of spiral MoTe2 to optimize collectively the above issues. The conductive atomic force microscopy of an individual spiral reveals that the retentive vertical conduction irrespective of layer thickness benefits from the connected screw dislocation lines between interlayers. Theoretical calculations uncover that the regions near the edge step of the spiral structures more easily form Te vacancies and have lower ΔGH * as extra active sites. A single spiral MoTe2 -based on-chip microcell was fabricated to extract HER activity and achieved an ultrahigh current density of 3000 mA cm-2 at an overpotential of 0.4 V, which is about two orders of magnitude higher than the exfoliated counterpart. Profoundly, this unusual spiral model will initiate a new pathway for triggering other inert catalytic reactions.

On-Chip Purification of Tetracyclines Based on Copper Ions Interaction.

Antibiotics are widely used to both prevent and treat bacterial diseases as well as promote animal growth. This massive use leads to the presence of residual antibiotics in food with severe consequences for human health. Limitations and regulations on the tolerated amount of antibiotics in food have been introduced and analytical methods have been developed. The bioanalytical methods usually employed to detect antibiotic residues, however, are time-consuming, expensive and laboratory-based. Novel methods with improved rapidity, portability and cost that are easy-to-use and sustainable are therefore highly desirable. In the attempt to fulfill this need, a microfluidic system was set up herein for the purification and pre-concentration of tetracyclines from raw milk selected as the case-study. The system includes a polymeric microfluidic chip containing magnetic beads loaded with copper to exploit the preferential interaction of tetracycline with divalent ions. The microfluidic system was demonstrated to efficiently pre-concentrate tetracycline, oxytetracycline and chlortetracycline with similar performances and efficiently purify tetracycline from raw milk without any pre-treatment. The simplified method described in this paper could be easily integrated in a compact and portable device for the in-field detection of tetracyclines, with the economic advantage of preventing food wastes and guaranteeing food safety.

Micromechanical Force Sensor Using the Stress-Impedance Effect of Soft Magnetic FeCuNbSiB.

By using the stress-impedance (SI) effect of a soft magnetic amorphous FeCuNbSiB alloy, a micromachined force sensor was fabricated and characterized. The alloy was used as a sputtered thin film of 500 nm thickness. To clarify the SI effect in the used material as a thin film, its magnetic and mechanical properties were first investigated. The stress dependence of the magnetic permeability was shown to be caused by the used transducer effect. The sputtered thin film also exhibited a large yield strength of 983 GPa. Even though the fabrication technology for the device is very simple, characterization revealed a gauge factor (GF) of 756, which is several times larger than that achieved with conventional transducer effects, such as the piezoresistive effect. The fabricated device shows great application potential as a tactile sensor.

A Two-Photon Microimaging-Microdevice System for Four-Dimensional Imaging of Local Drug Delivery in Tissues.

Advances in the intratumor measurement of drug responses have included a pioneering biomedical microdevice for high throughput drug screening in vivo, which was further advanced by integrating a graded-index lens based two-dimensional fluorescence micro-endoscope to monitor tissue responses in situ across time. While the previous system provided a bulk measurement of both drug delivery and tissue response from a given region of the tumor, it was incapable of visualizing drug distribution and tissue responses in a three-dimensional (3D) way, thus missing the critical relationship between drug concentration and effect. Here we demonstrate a next-generation system that couples multiplexed intratumor drug release with continuous 3D spatial imaging of the tumor microenvironment via the integration of a miniaturized two-photon micro-endoscope. This enables optical sectioning within the live tissue microenvironment to effectively profile the entire tumor region adjacent to the microdevice across time. Using this novel microimaging-microdevice (MI-MD) system, we successfully demonstrated the four-dimensional imaging (3 spatial dimensions plus time) of local drug delivery in tissue phantom and tumors. Future studies include the use of the MI-MD system for monitoring of localized intra-tissue drug release and concurrent measurement of tissue responses in live organisms, with applications to study drug resistance due to nonuniform drug distribution in tumors, or immune cell responses to anti-cancer agents.

Regeneration of Assembled, Molecular-Motor-Based Bionanodevices.

The guided gliding of cytoskeletal filaments, driven by biomolecular motors on nano/microstructured chips, enables novel applications in biosensing and biocomputation. However, expensive and time-consuming chip production hampers the developments. It is therefore important to establish protocols to regenerate the chips, preferably without the need to dismantle the assembled microfluidic devices which contain the structured chips. We here describe a novel method toward this end. Specifically, we use the small, nonselective proteolytic enzyme, proteinase K to cleave all surface-adsorbed proteins, including myosin and kinesin motors. Subsequently, we apply a detergent (5% SDS or 0.05% Triton X100) to remove the protein remnants. After this procedure, fresh motor proteins and filaments can be added for new experiments. Both, silanized glass surfaces for actin-myosin motility and pure glass surfaces for microtubule-kinesin motility were repeatedly regenerated using this approach. Moreover, we demonstrate the applicability of the method for the regeneration of nano/microstructured silicon-based chips with selectively functionalized areas for supporting or suppressing gliding motility for both motor systems. The results substantiate the versatility and a promising broad use of the method for regenerating a wide range of protein-based nano/microdevices.

Cytoplasmic fusion between an enlarged embryonic stem cell and a somatic cell using a microtunnel device.

Based on a previous finding that fusion of a somatic cell with an embryonic stem (ES) cell reprogrammed the somatic cell, genes for reprogramming transcription factors were selected and induced pluripotent stem (iPS) cell technology was developed. The cell fusion itself produced a tetraploid cell. To avoid nuclear fusion, a method for cytoplasmic fusion using a microtunnel device was developed. However, the ES cell was too small for cell pairing at the device. Therefore, in the present study, ES cell enlargement was carried out with the colchicine derivative demecolcine (DC). DC induced enlargement of ES cells without loss of their stemness. When an enlarged ES cell was paired with a somatic cell in the microtunnel device, cytoplasmic fusion was observed. The present method may be useful for further development of reprogramming techniques for iPS cell preparation without gene transfection.

Electro-osmotic surface effects generation in an electrokinetic-based transport device: A comparison of RF and MW plasma generating sources.

It is a common practice in insulator-based dielectrophoretic separation to use and reuse PDMS-constructed microdevice for an extended period of time while performing biological and technical replicate experiments. This is usually done to rule out any effects of device variation on separation efficiency. Ensuring that all experimental conditions remain the same is critical to the conclusion that can be drawn from such repeated experiments. One important contributing factor to the flow of materials within the device is electro-osmotic velocity, which stems from the surface condition of the device construction materials. In this paper, we present an affordable microwave-based (MESA-Mgen) oxygen plasma cleaner developed for approximately less than $100 using readily obtainable parts from an average local hardware store with no specialized tools. This low-cost room-air microwave plasma generator was designed using an R-4055, 400 W, 2450 MHz half-pint household microwave oven (Sharp®) for exploring the possibility of sealing polydimethylsiloxane (PDMS) devices onto glass with minimal budgetary commitment. Microfluidic channels generated using MESA-Mgen were evaluated for their electro-osmotic velocities while factors including contact angles, storage-solvent, half-way hydrophobicity period were also explored with MESA-Mgen, and the results were compared to those obtained from the commercially available plasma cleaner (COM-PC). These outcomes revealed that the MESA-Mgen induced hydrophilicity and ensured leak-free sealing of PDMS substrates in a manner comparable with the COM-PC.

Make it simple: long-term stable gradient generation in a microfluidic microdevice.

Microfluidics-based gradient generators have been used for various biological applications, specifically chemotaxis in cell culture. However, the ability to generate and maintain long term gradients alongside the ability to quickly switch solutions is a challenge of the current microfabricated systems. In this study, a simple flow-driven microfluidic system was developed to achieve long-term stable concentration gradients. Computational modelling was performed to highlight the fluid dynamics as well as to verify the ability of maintaining stable gradients over 7 days. Numerical simulation was analysed to evaluate the static pressure, velocity magnitude and wall shear stress distribution in the chamber. A microdevice fabricated with polydimethylsiloxane (PDMS), using a standard soft lithography technique is presented. It consists of eight parallel microchannels (5 µm × 30 µm × 1,800 µm) linking source and sink chambers; a syringe pump drives fluid through the sink chamber, advection/diffusion from source to sink establishes a gradient. A gradient of a fluorescent dye was generated under the low flow control at 1-10 µl/h of a simple syringe pump equipped with a pulsation damper that was comparable to a pulseless microfluidic pump. Concentration gradients were formed in 1 h and stable from 2 h out to 5 days and consuming less than 1.0 ml of solution. This study focuses on a novel solution to achieve a long-term microfluidic gradient generator using simple engineering techniques of biomedical microdevices.

Fabrication of a polycarbonate microdevice and boronic acid-mediated surface modification for on-chip sample purification and amplification of foodborne pathogens.

In this study, we integrated sample purification and genetic amplification in a seamless polycarbonate microdevice to facilitate foodborne pathogen detection. The sample purification process was realized based on the increased affinity of the boronic acid-modified surface toward the cis-diol group present on the bacterial outer membrane. The modification procedure was conducted at room temperature using disposable syringe. The visible color and fluorescence signals of alizarin red sodium were used to confirm the success of the surface modification process. Escherichia coli O157:H7 containing green fluorescence protein (GFP) and Staphylococcus aureus were chosen as the microbial models to demonstrate the nonspecific immobilization using the microdevice. Bacterial solutions of various concentrations were injected into the microdevice at three flow rates to optimize the operation conditions. This microdevice successfully amplified the 384-bp fragment of the eaeA gene of the captured E. coli O157:H7 within 1 h. Its detection limit for E. coli O157:H7 was determined to be 1 × 103 colony-forming units per milliliter (CFU mL-1). The proposed microdevice serves as a monolithic platform for facile and on-site identification of major foodborne pathogens.

Hybrid elastomer-plastic microfluidic device as a convenient model for mimicking the blood-brain barrier in vitro.

In this study, we fabricated a hybrid elastomer-plastic microdevice using the silicone elastomer poly(dimethylsiloxane) (PDMS) and the plastic polycarbonate (PC), to mimic the human blood-brain barrier (BBB) in vitro. Specifically, the microchannel-imprinted elastomer was first coated with 3-aminopropyltriethoxysilane to produce amine-terminated PDMS. Then, simply by conformal contact at room temperature, the amine-functionalized PDMS was bonded to pristine PC through the formation of urethane linkages. Aside from realizing device bonding, the amine functionalization also assisted in subsequent dopamine coating to form polydopamine and provide a stable surface for culturing human endothelial cells and central nervous system-related cells (e.g., astrocytes) inside the microchannels. Successful mimicking of the BBB-like microenvironment was assessed by 3D co-culturing of human endothelial cells and astrocytes, where the microdevice was verified as an acceptable in vitro BBB model according to the following four criteria: the formation of tight junctions at the cell-cell boundaries of the endothelial cells, evaluated by the expression of the tight junction marker ZO-1; the formation of actin filaments, evaluated using rhodamine phalloidin dye; low permeability, tested using the fluorescent tracer 40-kDa FITC-dextran; and good transendothelial electrical resistance (a measure of the tight junction integrity formed between the endothelial cells). The fabricated PDMS-PC microfluidic device ensured simple yet stable device sealing, and simultaneously enhanced BBB-mimicking cell attachment, thus fulfilling all major criteria for its application as a convenient in vitro BBB model.

Evaporation-induced stimulation of bacterial osmoregulation for electrical assessment of cell viability.

Bacteria cells use osmoregulatory proteins as emergency valves to respond to changes in the osmotic pressure of their external environment. The existence of these emergency valves has been known since the 1960s, but they have never been used as the basis of a viability assay to tell dead bacteria cells apart from live ones. In this paper, we show that osmoregulation provides a much faster, label-free assessment of cell viability compared with traditional approaches that rely on cell multiplication (growth) to reach a detectable threshold. The cells are confined in an evaporating droplet that serves as a dynamic microenvironment. Evaporation-induced increase in ionic concentration is reflected in a proportional increase of the droplet's osmotic pressure, which in turn, stimulates the osmoregulatory response from the cells. By monitoring the time-varying electrical conductance of evaporating droplets, bacterial cells are identified within a few minutes compared with several hours in growth-based methods. To show the versatility of the proposed method, we show detection of WT and genetically modified nonhalotolerant cells (Salmonella typhimurium) and dead vs. live differentiation of nonhalotolerant (such as Escherichia coli DH5α) and halotolerant cells (such as Staphylococcus epidermidis). Unlike the growth-based techniques, the assay time of the proposed method is independent of cell concentration or the bacteria type. The proposed label-free approach paves the road toward realization of a new class of real time, array-formatted electrical sensors compatible with droplet microfluidics for laboratory on a chip applications.