Rapid Electrohydrodynamic-Driven Pattern Replication over a Large Area via Ultrahigh Voltage Pulses.

Despite the prospects of electrohydrodynamic instability patterning (EHIP), poor process parameter controllability is a significant challenge in uniform large-scale nanopatterning. Herein, we introduce a EHIP process using an ultrahigh electric field (>108 V/m) to effectively accelerate the pattern growth evolution. Owing to the strong dependence on a temporal parameter (1/τm) of the field strength, our method not only reduces the completion time of pattern growth but also overcomes critical parametric restrictions on the pattern replication, thereby enhancing the replicated pattern quality in three dimensions. The pattern can be uniformly replicated over the entire film surface even without a perfectly uniform air gap, which has been severely difficult in the conventional method. To further demonstrate how straightforward yet versatile our approach is, we applied our EHIP approach to successfully replicate the densely packed nanostructures of cicada wings.

Conductive Thread-Based Immunosensor for Pandemic Influenza A (H1N1) Virus Detection.

Infectious agents such as viruses pose significant threats to human health, being transmitted via direct contact as well as airborne transmission without direct contact, thus requiring rapid detection to prevent the spread of infectious diseases. In this study, we developed a conductive thread-based immunosensor (CT-IS), a biosensor to easily detect the presence of airborne viruses. CT-IS utilizes an antibody that specifically recognizes the HA protein of the pandemic influenza A (pH1N1) virus, which is incorporated into the conductive thread. The antigen-antibody interaction results in increased strain on the conductive thread in the presence of the pH1N1 virus, resulting in increased electrical resistance of the CT-IS. We evaluated the performance of this sensor using the HA protein and the pH1N1 virus, in addition to samples from patients infected with the pH1N1 virus. We observed a significant change in resistance in the pH1N1-infected patient samples (positive: n = 11, negative: n = 9), whereas negligible change was observed in the control samples (patients not infected with the pH1N1 virus; negative). Hence, the CT-IS is a lightweight fiber-type sensor that can be used as a wearable biosensor by combining it with textiles, to detect the pH1N1 virus in a person's vicinity.

Real-time and label-free biosensing using moiré pattern generated by bioresponsive hydrogel.

Bioresponsive hydrogels are smart materials that respond to various external stimuli and exhibit great potential as biosensors owing to their capability of real-time and label-free detection. Here, we propose a sensing platform based on bioresponsive hydrogels, employing the concept of moiré patterns. Two sets of line patterns with different pitch sizes are prepared; a hydrogel grating whose pitch size changes according to external stimuli and a reference grating with constant pitch size. The volume changes of the hydrogel caused by external stimuli changes the pitch size of the hydrogel grating, and subsequently, the pitch sizes of the moiré patterns (moiré signal), whose values can be obtained in a real-time and label-free manner through customized moiré microscopy and signal processing. After confirming that the pH-induced swelling of hydrogel could be monitored using moiré patterns, we performed moiré pattern-based detection of specific proteins using protein-responsive hydrogel that underwent shrinking via interaction with target proteins. Brain-derived neurotrophic factor and platelet-derived growth factor were selected as the model proteins, and our proposed system successfully detected both proteins at nanomolar levels. In both cases, the pitch size change of hydrogel grating was monitored much more sensitively using moiré patterns than through direct measurements. The changes in the moiré signals caused by target proteins were detected in ex-vivo environments using a custom-made intraocular lens incorporating the hydrogel grating, demonstrating the capability of the proposed system to detect various markers in intraocular aqueous humor, when implanted in the eye.

High diopter spectacle using a flexible Fresnel lens with a combination of grooves.

In this study, the Fresnel lens was investigated as a potential candidate for vision correction in patients with myopia. A few previous studies have suggested this idea; however, Fresnel lenses are limited by their aesthetics and quality. Therefore, we designed a combination of Fresnel lens grooves with a constant height and pitch of 13 µm and 0.1 mm, respectively, to overcome the limitations caused by ultra-precision machining with a tool nose radius of 30 µm. A thin replicated Fresnel lens with a power of -5 diopter was procured and applied directly as spectacles that are unattached to the normal lens. The optical performance and image quality of the Fresnel lens were compared with those of a conventional lens possessing the same power in both near and far vision. These results extend the applicability for the use of Fresnel lenses as vision-correcting ophthalmological lenses and imaging systems.

Fabrication of Nanostructures on a Large-Area Substrate with a Minimized Stitch Error Using the Step-and-Repeat Nanoimprint Process.

Nanoimprint lithography (NIL) is suitable for achieving high uniformity and mass production. However, in conventional NIL, a stamp suitable for the substrate size is required to increase the substrate size. To address this issue, we fabricated nanostructures on a large-area substrate using step-and-repeat NIL after making a small stamp. A stamp was produced using glass, and a nano-pillar pattern with a diameter of 600 nm, an interval of 400 nm, and a height of 270 nm was used during the experiment. The area of the pattern on the stamp was 10 mm × 10 mm, and the step-and-repeat process was performed 25 times to transfer the nanostructures to a 4-inch substrate. In addition, stitch gaps were created between the patterns, which could decrease the performance upon future application. To minimize this stitch gap, a high-precision glass scale was attached to the stamp feeder to precisely control the position and to minimize the step difference. Moreover, an experiment was conducted to minimize the stitch gap by adjusting the movement interval of the stamp, and the stitch spacing was minimized by moving the stamp position by 9.97 mm. This approach will facilitate the manufacturing of large-area substrates and other structures in the future.

Microfluidic device for one-step detection of breast cancer-derived exosomal mRNA in blood using signal-amplifiable 3D nanostructure.

Metastasis attributed to approximately 90% of cancer-related deaths; hence, the detection of metastatic tumor-derived components in the blood assists in determining cancer recurrence and patient survival. Microfluidic-based sensors facilitate analysis of small fluid volumes and represent an accurate, rapid, and user-friendly method of field diagnoses. In this study, we have developed a microfluidic chip-based exosomal mRNA sensor (exoNA-sensing chip) for the one-step detection of exosomal ERBB2 in the blood by integrating a microfluidic chip and 3D-nanostructured hydrogels. The exoNA-sensing chip is a vacuum-driven power-free microfluidic chip that can accurately control the flow of trace fluids (<100 µL). The sensing part of the exoNA-sensing chip includes 3D-nanostructured hydrogels capable of detecting ERBB2 and a reference gene by amplifying a fluorescent signal via an enzyme-free catalytic hairpin assembly reaction at room temperature. This hydrogel offers a detection limit of 58.3 fM with good selectivity for target sequences. The performance of the exoNA-sensing chip was evaluated by testing in vitro and in vivo samples and was proven to be effective for cancer diagnosis and liquid biopsies.

Hierarchically Porous, Laser-Pyrolyzed Carbon Electrode from Black Photoresist for On-Chip Microsupercapacitors.

We report a laser-pyrolyzed carbon (LPC) electrode prepared from a black photoresist for an on-chip microsupercapacitor (MSC). An interdigitated LPC electrode was fabricated by direct laser writing using a high-power carbon dioxide (CO2) laser to simultaneously carbonize and pattern a spin-coated black SU-8 film. Due to the high absorption of carbon blacks in black SU-8, the laser-irradiated SU-8 surface was directly exfoliated and carbonized by a fast photo-thermal reaction. Facile laser pyrolysis of black SU-8 provides a hierarchically macroporous, graphitic carbon structure with fewer defects (ID/IG = 0.19). The experimental conditions of CO2 direct laser writing were optimized to fabricate high-quality LPCs for MSC electrodes with low sheet resistance and good porosity. A typical MSC based on an LPC electrode showed a large areal capacitance of 1.26 mF cm-2 at a scan rate of 5 mV/s, outperforming most MSCs based on thermally pyrolyzed carbon. In addition, the results revealed that the high-resolution electrode pattern in the same footprint as that of the LPC-MSCs significantly affected the rate performance of the MSCs. Consequently, the proposed laser pyrolysis technique using black SU-8 provided simple and facile fabrication of porous, graphitic carbon electrodes for high-performance on-chip MSCs without high-temperature thermal pyrolysis.

Highly Sensitive and Reliable microRNA Detection with a Recyclable Microfluidic Device and an Easily Assembled SERS Substrate.

Surface-enhanced Raman spectroscopy (SERS) detection in microfluidics is an interesting topic because of its high sensitivity, miniaturization, and ability to perform online detection. However, the difficulties in generating SERS-based microfluidic devices with uniform signal reproducibility and high sensitivity have hindered their widespread application. In addition, the recyclability of the SERS-based microfluidic devices can contribute to their broad commercialization, but the possible contamination in the detection area and cumbersome cleaning procedures remain a challenge. In this study, we describe a repeatable SERS-based microfluidic device comprising a disposable SERS substrate and a reusable microfluidic channel. The microfluidic channel was prepared via mechanical processing, and the SERS substrate was fabricated by nanoimprint lithography and electrodeposition. The SERS substrate and microfluidic channel can be attached easily because they were assembled using screws. The SERS substrate achieved an excellent SERS enhancement factor greater than 108 over a large sample area, signal uniformity, and substrate-to-substrate reproducibility. This guaranteed reliable and sensitive signals in every experiment. Furthermore, the disposable SERS substrate contributed exact detection of target molecules. Finally, their practical application was demonstrated with the repeated use of the microfluidic device by detecting a specific micro-RNA, (miR-34a) at a concentration as low as 5 fM.

Biomimetic Nanopillar-Based Biosensor for Label-Free Detection of Influenza A Virus.

Since the first emergence of influenza viruses, they have caused the flu seasonally worldwide. Precise detection of influenza viruses is required to prevent the spreading of the disease. Herein, we developed an optical biosensor using peptide-immobilized nanopillar structures for the label-free detection of influenza viruses. The spin-on-glass nanopillar structures were fabricated by nanoimprint lithography. A sialic acid-mimic peptide, which can specifically bind to hemagglutinin on the surface of the influenza virus, was immobilized onto the nanopillars via polymerized dopamine. The constructed nanopillar sensor enabled us to detect influenza A viruses in the range of 103-105 plaque-forming units through simple measurements of reflectance. Our findings suggest that biomimetic modification of nanopillar structures can be an alternative method for the immunodiagnosis of influenza viruses.

Fog Collection Based on Secondary Electrohydrodynamic-Induced Hybrid Structures with Anisotropic Hydrophilicity.

The outcomes of the study of plant surfaces, such as rice leaves or bamboo leaves, have led to extensive efforts being devoted to fabricating anisotropic arrays of micro/nanoscale features for exploring anisotropic droplet spreading. Nonetheless, precise engineering of the density and continuity of three-phase contact lines for anisotropic wetting remains a significant challenge without resorting to chemical modifications and costly procedures. In this work, we investigated secondary electrohydrodynamic instability in polymer films for producing secondary nanosized patterns between the micrometer-sized grooves by controlling the timescale parameter, 1/τm (>10-4 s-1). We experimentally demonstrated facile morphological control of anisotropic wettability without the use of any chemical modifications. Thus, anisotropic hydrophilic surfaces fabricated by the secondary phase instability of polymer films are advantageous for both droplet condensation and removal, thereby outperforming the water collection efficiency of conventional (isotropic) hydrophilic surfaces in water harvesting applications (∼200 mg·cm-2·h-1) with excellent durability.

Three-Dimensional Printable Hydrogel Using a Hyaluronic Acid/Sodium Alginate Bio-Ink.

Bio-ink properties have been extensively studied for use in the three-dimensional (3D) bio-printing process for tissue engineering applications. In this study, we developed a method to synthesize bio-ink using hyaluronic acid (HA) and sodium alginate (SA) without employing the chemical crosslinking agents of HA to 30% (w/v). Furthermore, we evaluated the properties of the obtained bio-inks to gauge their suitability in bio-printing, primarily focusing on their viscosity, printability, and shrinkage properties. Furthermore, the bio-ink encapsulating the cells (NIH3T3 fibroblast cell line) was characterized using a live/dead assay and WST-1 to assess the biocompatibility. It was inferred from the results that the blended hydrogel was successfully printed for all groups with viscosities of 883 Pa∙s (HA, 0% w/v), 1211 Pa∙s (HA, 10% w/v), and 1525 Pa∙s, (HA, 30% w/v) at a 0.1 s-1 shear rate. Their structures exhibited no significant shrinkage after CaCl2 crosslinking and maintained their integrity during the culture periods. The relative proliferation rate of the encapsulated cells in the HA/SA blended bio-ink was 70% higher than the SA-only bio-ink after the fourth day. These results suggest that the 3D printable HA/SA hydrogel could be used as the bio-ink for tissue engineering applications.

Integrated, Automated, Fast PCR System for Point-Of-Care Molecular Diagnosis of Bacterial Infection.

We developed an integrated PCR system that performs automated sample preparation and fast polymerase chain reaction (PCR) for application in point-of care (POC) testing. This system is assembled from inexpensive 3D-printing parts, off-the-shelf electronics and motors. Molecular detection requires a series of procedures including sample preparation, amplification, and fluorescence intensity analysis. The system can perform automated DNA sample preparation (extraction, separation and purification) in ≤5 min. The variance of the automated sample preparation was clearly lower than that achieved using manual DNA extraction. Fast thermal ramp cycles were generated by a customized thermocycler designed to automatically transport samples between heating and cooling blocks. Despite the large sample volume (50 µL), rapid two-step PCR amplification completed 40 cycles in ≤13.8 min. Variations in fluorescence intensity were measured by analyzing fluorescence images. As proof of concept of this system, we demonstrated the rapid DNA detection of pathogenic bacteria. We also compared the sensitivity of this system with that of a commercial device during the automated extraction and fast PCR of Salmonella bacteria.

Parametric scheme for rapid nanopattern replication via electrohydrodynamic instability.

Electrohydrodynamic (EHD) instability patterning exhibits substantial potential for application as a next-generation lithographic technique; nevertheless, its development continues to be hindered by the lack of process parameter controllability, especially when replicating sub-microscale pattern features. In this paper, a new parametric guide is introduced. It features an expanded range of valid parameters by increasing the pattern growth velocity, thereby facilitating reproducible EHD-driven patterning for perfect nanopattern replication. Compared with conventional EHD-driven patterning, the rapid patterning approach not only shortens the patterning time but also exhibits enhanced scalability for replicating small and geometrically diverse features. Numerical analyses and simulations are performed to elucidate the interplay between the pattern growth velocity, fidelity of the replicated features, and boundary between the domains of suitable and unsuitable parametric conditions in EHD-driven patterning. The developed rapid route facilitates nanopattern replication using EHD instability with a wide range of suitable parameters and further opens up many opportunities for device applications using tailor-made nanostructures in an effective and straightforward manner.

Selective Transfer of Light-Emitting Diodes onto a Flexible Substrate via Laser Lissajous Scanning.

As the light-emitting diode (LED) size gradually decreases, it is difficult to conventionally transfer an LED onto a donor substrate. In this paper, we propose a print transfer method that selectively transfers an LED onto a UV release tape, i.e., the donor substrate, via focused laser scanning with Lissajous patterns. We implemented an optical system based on focused laser scanning to perform selective transfer; this can adjust the scanning area immediately without changing the donor substrate size according to the LED size. Because the commercialized UV release tape is utilized as a donor substrate, the adhesion between the LED and donor substrate can be constantly maintained even after repeated experiments. In this study, several LEDs were transferred to a flexible printed circuit board-arranged in a circular and square shape to demonstrate a high degree of freedom of the system-and turned on.

Peptidoglycan-Binding Protein Metamaterials Mediated Enhanced and Selective Capturing of Gram-Positive Bacteria and Their Specific, Ultra-Sensitive, and Reproducible Detection via Surface-Enhanced Raman Scattering.

Biological metamaterials with a specific size and spacing are necessary for developing highly sensitive and selective sensing systems to detect hazardous bacteria in complex solutions. Herein, the construction of peptidoglycan-binding protein (PGBP)-based metamaterials to selectively capture Gram-positive cells with high efficacy is reported. Nanoimprint lithography was used to generate a nanohole pattern as a template, the inside of which was modified with nickel(II)-nitrilotriacetic acid (Ni-NTA). Then, PGBP metamaterials were fabricated by immobilizing PGBP via chelation between Ni-NTA and six histidines on PGBP. Compared to the flat and spread PGBP-covered bare substrates, the PGBP-based metamaterials enabled selective capturing of Gram-positive bacteria with high efficacy, owing to enhanced interactions between the metamaterials and bacterial surface not shown in bulk materials. Thereafter, the specific strain and quantitative information of the captured bacteria was obtained by surface-enhanced Raman scattering mapping analysis in the 1 to 1 × 106 cfu/mL range within 30 min. It should be noted that no additional signal amplification process was required for lowly abundant bacteria, even at the single-bacterium level. The PGBP-based metamaterials could be regenerated multiple times with preserved sensing efficiency. Finally, this assay can detect specific Gram-positive bacteria, such as Staphylococcus aureus, in human plasma.

A Lipid-Nanopillar-Array-Based Immunosorbent Assay.

Since infectious diseases, particularly viral infections, have threatened human health and caused huge economical losses globally, a rapid, sensitive, and selective virus detection platform is highly demanded. Enzyme-linked immunosorbent assay (ELISA) with flat solid substrates has been dominantly used in detecting whole viruses for its straightforwardness and simplicity in assay protocols, but it often suffers from limited sensitivity, poor quantification range, and a time-consuming assay procedure. Here, a lipid-nanopillar-array-based immunosorbent assay (LNAIA) is developed with a nanopillar-supported lipid bilayer substrate with fluorophore-modified antibodies for rapid, sensitive, and quantitative detection of viruses. 3D nanopillar array structures and fluid antibodies with fluorophores facilitate faster and efficient target binding and rapid fluorophore localization for quick, reliable analysis on binding events with a conventional fluorescence microscopy setup. LNAIA enables quantification of H1N1 virus that targets down to 150 virus particles with 5-orders-of-magnitude dynamic range within 25 min, which cannot be achieved with conventional ELISA platforms.

System for Fabrication of Large-Area Roll Molds by Step-and-Repeat Liquid Transfer Imprint Lithography.

The effective production of nanopatterned films generally requires a nanopatterned roll mold with a large area. We report on a novel system to fabricate large-area roll molds by recombination of smaller patterned areas in a step-and-repeat imprint lithography process. The process is accomplished in a method similar to liquid transfer imprint lithography (LTIL). The stamp roll with a smaller area takes up the liquid resist by splitting from a donor substrate or a donor roll. The resist is then transferred from a stamp roll to an acceptor roll and stitched together in a longitudinal and, if necessary, in a circumferential direction. During transfer, the nanostructured resist is UV-exposed and crosslinked directly on the acceptor roll. The acceptor roll with the stitched and recombined stamp patterns is ready to be used as a large-area roll mold for roll-based imprinting. A system for this purpose was designed, and its operation was demonstrated taking the example of an acceptor roll of 1 m length and 250 mm diameter, which was covered by 56 patterned areas. Such a system represents an elegant and efficient tool to recombine small patterned areas directly on a large roll mold and opens the way for large-area roll-based processing.

An electrophoretic DNA extraction device using a nanofilter for molecular diagnosis of pathogens.

Rapid and efficient nucleic acid (NA) extraction and concentration are required for point-of-care analysis in order to prevent an epidemic/pandemic disease outbreak. Typical silica-based NA extraction methods have limitations such as being time-consuming, requiring human intervention, and resulting in a low recovery yield. In this study, we have developed a pathogenic DNA extraction device based on electrokinetic separation incorporated with a silicon nitride (SiNx) nanofilter, which expedites the DNA extraction procedure with advantages of being convenient, efficient, and inexpensive. This DNA extraction device consists of a computer numerical control (CNC) milled-Teflon gadget with a cis-chamber as a cell lysate reservoir and a trans-chamber as a elution solution reservoir, with the SiNx nanofilter being inserted between the two chambers. The SiNx nanofilter was fabricated using a photolithographic method in conjunction with nanoimprinting. Approximately 7.2 million nanopores of 220 nm diameter were located at the center of the nanofilter. When a DC electric field is applied through the nanopores, DNA is transferred from the cis-chamber to the trans-chamber to isolate the DNA from the cell debris. To demonstrate the DNA extraction performance, we measured the absorbances at 260 and 280 nm and performed a real-time polymerase chain reaction (real-time PCR) using the recovered DNA to verify its feasibility for downstream genetic analysis. Moreover, the DNA extraction device was successfully operated using a 1.5 V alkaline battery, which verifies the portability of the device for point-of-care testing. Such an advanced DNA extraction system can be utilized in various fields including clinical analysis, pathogen detection, forensic analysis, and on-site detection.

Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane.

A method to directly collect negatively charged nucleic acids, such as DNA and RNA, in the biosamples simply by applying an electric field in between the sample and collection buffer separated by the nanofilter membrane is proposed. The nanofilter membrane was made of low-stress silicon nitride with a thickness of 100 nm, and multiple pores were perforated in a highly arranged pattern using nanoimprint technology with a pore size of 200 nm and a pore density of 7.22 × 108/cm2. The electrophoretic transport of hsa-mir-93-5p across the membrane was confirmed in pure microRNA (miRNA) mimic solution using quantitative reverse transcription-polymerase chain reactions (qRT-PCR). Consistency of the collected miRNA quantity, stability of the system during the experiment, and yield and purity of the prepared sample were discussed in detail to validate the effectiveness of the electrical protocol. Finally, in order to check the applicability of this method to clinical samples, liquid biopsy process was demonstrated by evaluating the miRNA levels in sera of hepatocellular carcinoma patients and healthy controls. This efficient system proposed a simple, physical idea in preparation of nucleic acid from biosamples, and demonstrated its compatibility to biological downstream applications such as qRT-PCR as the conventional nucleic acid extraction protocols.

Fabrication and Characterization of Bioresorbable Drug-coated Porous Scaffolds for Vascular Tissue Engineering.

Bioresorbable polymers have been studied for several decades as attractive candidates for promoting the advancement of medical science and bio-technology in modern society. In particular, with a well-defined architecture, bioresorbable polymers have prominent advantages over their bulk counterparts for applications in biomedical and implant devices, such as cell delivery, scaffolds for tissue engineering, and hydrogels as well as in the pharmaceutical fields. Biocompatible implant devices based on bioresorbable materials (for instance, bioresorbable polymers that combine the unique advantages of biocompability and easy handling) have emerged as a highly active field due to their promising applications in artificial implant systems and biomedical devices. In this paper, we report an approach to fabricate porous polycaprolactone (PCL) scaffolds using a 3D printing system. And its surface was treated to a hydrophilic surface using plasma treatment. Then, the aspirin and atorvastatin calcium salt mixture was dip coated onto the surface. The drug coating technology was used to deposit the drug material onto the scaffold surface. Our porous PCL scaffold was coated with aspirin and atorvastatin calcium salt to reduce the blood LDL cholesterol and restenosis. These results suggest that our approach may provide a promising scaffold for developing bioresorbable drug-delivery-biomaterials. We further demonstrate that our bioresorbable medical device can be used as vascular scaffolds to provide a wide range of applications for the design of medical devices.