Bioengineered amyloid peptide for rapid screening of inhibitors against main protease of SARS-CoV-2.

The coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has evoked a worldwide pandemic. As the emergence of variants has hampered the neutralization capacity of currently available vaccines, developing effective antiviral therapeutics against SARS-CoV-2 and its variants becomes a significant challenge. The main protease (Mpro) of SARS-CoV-2 has received increased attention as an attractive pharmaceutical target because of its pivotal role in viral replication and proliferation. Here, we generated a de novo Mpro-inhibitor screening platform to evaluate the efficacies of Mpro inhibitors based on Mpro cleavage site-embedded amyloid peptide (MCAP)-coated gold nanoparticles (MCAP-AuNPs). We fabricated MCAPs comprising an amyloid-forming sequence and Mpro-cleavage sequence, mimicking in vivo viral replication process mediated by Mpro. By measuring the proteolytic activity of Mpro and the inhibitory efficacies of various drugs, we confirmed that the MCAP-AuNP-based platform was suitable for rapid screening potential of Mpro inhibitors. These results demonstrated that our MCAP-AuNP-based platform has great potential for discovering Mpro inhibitors and may accelerate the development of therapeutics against COVID-19.

Plasma-based diagnostic and screening platform using a combination of biosensing signals in Alzheimer's disease.

Using biosensor to screen for Alzheimer's disease (AD) facilitates early detection of AD with high sensitivity and accuracy. This approach overcomes the limitations of conventional AD diagnostic methods, such as neuropsychological assessment and neuroimaging analysis. Here, we propose a simultaneous analysis of signal combinations generated by four crucial AD biomarkers (Amyloid beta 1-40 (Aß40), Aß42, total tau 441 (tTau441), and phosphorylated tau 181 (pTau181)) by inducing a dielectrophoretic (DEP) force on fabricated interdigitated microelectrode (IME) sensor. By applying an optimal DEP force, our biosensor selectively concentrates and filters the plasma-based AD biomarkers, exhibiting high sensitivity (limit of detection <100 fM) and selectivity in the plasma-based AD biomarkers detection (p < 0.0001). Consequently, it is demonstrated that a complex combined signal comprising four AD-specific biomarker signals (Aß40- Aß42+ tTau441- pTau181) can differentiate between patients with AD and healthy subjects with high accuracy (78.85%) and precision (80.95%) (p < 0.0001).

Caco-2 cell-derived biomimetic electrochemical biosensor for cholera toxin detection.

Cholera is a highly contagious and lethal waterborne disease induced by an infection with Vibrio cholerae (V. cholerae) secreting cholera toxin (CTx). Cholera toxin subunit B (CTxB) from the CTx specifically binds with monosialo-tetra-hexosyl-ganglioside (GM1) found on the exterior cell membrane of an enterocyte. Bioinspired by the pathological process of CTx, we developed an electrochemical biosensor with GM1-expressing Caco-2 cell membrane (CCM) on the electrode surface. Briefly, the electrode surface was functionalized with CCM using the vesicle fusion method. We determined the CTxB detection performances of Caco-2 cell membrane-coated biosensor (CCB) using electrochemical impedance spectroscopy (EIS). the CCB had an excellent limit of detection of ∼11.46 nM and a detection range spanning 100 ng/mL - 1 mg/mL. In addition, the CCB showed high selectivity against various interfering molecules, including abundant constituents of intestinal fluid and various bacterial toxins. The long-term stability of the CCBs was also verified for 3 weeks using EIS. Overall, the CCB has excellent potential for practical use such as point-of-care and cost-effective testing for CTxB detection in developing countries.

Observation of electronic modes in open cavity resonator.

The resemblance between electrons and optical waves has strongly driven the advancement of mesoscopic physics, evidenced by the widespread use of terms such as fermion or electron optics. However, electron waves have yet to be understood in open cavity structures which have provided contemporary optics with rich insight towards non-Hermitian systems and complex interactions between resonance modes. Here, we report the realization of an open cavity resonator in a two-dimensional electronic system. We studied the resonant electron modes within the cavity and resolved the signatures of longitudinal and transverse quantization, showing that the modes are robust despite the cavity being highly coupled to the open background continuum. The transverse modes were investigated by applying a controlled deformation to the cavity, and their spatial distributions were further analyzed using magnetoconductance measurements and numerical simulation. These results lay the groundwork to exploring matter waves in the context of modern optical frameworks.

Biomimetically Engineered Amyloid-Shelled Gold Nanocomplexes for Discovering α-Synuclein Oligomer-Degrading Drugs.

The assembly of α-synuclein (αS) oligomers is recognized as the main pathological driver of synucleinopathies. While the elimination of toxic αS oligomers shows promise for the treatment of Parkinson's disease (PD), the discovery of αS oligomer degradation drugs has been hindered by the lack of proper drug screening tools. Here, we report a drug screening platform for monitoring the efficacy of αS-oligomer-degrading drugs using amyloid-shelled gold nanocomplexes (ASGNs). We fabricate ASGNs in the presence of dopamine, mimicking the in vivo generation process of pathological αS oligomers. To test our platform, the first of its kind for PD drugs, we use αS-degrading proteases and various small molecular substances that have shown efficacy in PD treatment. We demonstrate that the ASGN-based in vitro platform has strong potential to discover effective αS-oligomer-targeting drugs, and thus it may reduce the attrition problem in drug discovery for PD treatment.

One-Lead Single-Electron Source with Charging Energy.

Single-electron sources, formed by a quantum dot (QD), are key elements for realizing electron analogue of quantum optics. We develop a new type of single-electron source with functionalities that are absent in existing sources. This source couples with only one lead. By an AC rf drive, it successively emits holes and electrons cotraveling in the lead, as in the mesoscopic capacitor. Thanks to the considerable charging energy of the QD, however, emitted electrons have energy levels a few tens of millielectronvolts above the Fermi level, so that emitted holes and electrons are split by a potential barrier on demand, resulting in a rectified quantized current. The resulting pump map exhibits quantized triangular islands, in good agreement with our theory. We also demonstrate that the source can be operated with another tunable-barrier single-electron source in a series double QD geometry, showing parallel electron pumping by a common gate driving.

Electric-Field-Mediated In-Sensor Alignment of Antibody's Orientation to Enhance the Antibody-Antigen Binding for Ultrahigh Sensitivity Sensors.

Applying an electric-field (E-field) during antibody immobilization aligns the orientation of the antibody on the biosensor surface, thereby enhancing the binding probability between the antibody and antigen and maximizing the sensitivity of the biosensor. In this study, a biosensor with enhanced antibody-antigen binding probability was developed using the alignment of polar antibodies (immunoglobulin G [IgG]) under an E-field applied inside the interdigitated electrodes. The optimal alignment condition was first theoretically calculated and then experimentally confirmed by comparing the impedance change before and after the alignment of IgG (a purified anti-ß-amyloid antibody). With the optimized condition, the impedance change of the biosensor was maximized because of the alignment of IgG orientation on the sensor surface; the detection sensitivity of the antigen amyloid-beta 1-42 was also maximized. The E-field-based in-sensor alignment of antibodies is an easy and effective method for enhancing biosensor sensitivity.

Scalable Functionalization of Polyaniline-Grafted rGO Field-Effect Transistors for a Highly Sensitive Enzymatic Acetylcholine Biosensor.

For decades, acetylcholine (Ach) has been considered a critical biomarker for several degenerative brain diseases, including Alzheimer's, Parkinson's disease, Huntington's disease, and schizophrenia. Here, we propose a wafer-scale fabrication of polyaniline (PAni)-grafted graphene-based field-effect transistors (PGFET) and their biosensing applications for highly sensitive and reliable real-time monitoring of Ach in flow configuration. The grafted PAni provides suitable electrostatic binding sites for enzyme immobilization and enhances the pH sensitivity (2.68%/pH), compared to that of bare graphene-FET (1.81%/pH) for a pH range of 3-9 without any pH-hysteresis. We further evaluated the PGFET's sensing performance for Ach detection with a limit of detection at the nanomolar level and significantly improved sensitivity (~103%) in the concentration range of 108 nM to 2 mM. Moreover, the PGFET exhibits excellent selectivity against various interferences, including glucose, ascorbic acid, and neurotransmitters dopamine and serotonin. Finally, we investigated the effects of an inhibitor (rivastigmine) on the AchE activity of the PGFET. From the results, we demonstrated that the PGFET has great potential as a real-time drug-screening platform by monitoring the inhibitory effects on enzymatic activity.

Screening for cerebral amyloid angiopathy based on serological biomarkers analysis using a dielectrophoretic force-driven biosensor platform.

We aimed to analyze plasma amyloid-ß (Aß)1-40 and Aß1-42 using a highly sensitive dielectrophoretic-driven biosensor platform to demonstrate the possibility of precise cerebral amyloid angiopathy (CAA) diagnosis in participants classified according to Aß-positron emission tomography (PET) positivity and the neuroimaging criteria for CAA. We prospectively recruited 25 people with non-Alzheimer's disease (non-AD) and 19 patients with Alzheimer's disease (AD), which were further classified into the CAA- and CAA+ (possible and probable CAA) groups according to the modified Boston criteria. Patients underwent plasma Aß analysis using a highly sensitive nano-biosensor platform, Aß-PET scanning, and detailed neuropsychological testing. As a result, the average signal levels of Aß1-42/1-40 differed significantly between the non-AD and AD groups, and the CAA+ group exhibited significantly higher Aß1-40 signal levels than the CAA- group in both non-AD and AD groups. The concordance between the Aß1-40 signal level and the neuroimaging criteria for CAA was nearly perfect, with areas under the curve of 0.954 (95% confidence interval (CI) 0.856-1.000), 0.969 (0.894-1.000), 0.867 (0.648-1.000), and 1.000 (1.000-1.000) in the non-AD/CAA- vs. non-AD/possible CAA, non-AD/CAA- vs. non-AD/probable CAA, AD/CAA- vs. AD/possible CAA, and AD/CAA- vs. AD/probable CAA groups, respectively. Higher Aß1-40 signal levels were significantly associated with the presence of CAA according to regression analyses, and the neuroimaging pattern analysis partly supported this result. Our findings suggest that measuring plasma Aß1-40 signal levels using a highly sensitive biosensor platform could be a useful non-invasive CAA diagnostic method.

Technological advances in electrochemical biosensors for the detection of disease biomarkers.

With an increasing focus on health in contemporary society, interest in the diagnosis, treatment, and prevention of diseases has grown rapidly. Accordingly, the demand for biosensors for the early diagnosis of disease is increasing. However, the measurement range of existing electrochemical sensors is relatively high, which is not suitable for early disease diagnosis, requiring the detection of small amounts of biocomponents. Various attempts have been made to overcome this and amplify the signal, including binding with various labeling molecules, such as DNA, enzymes, nanoparticles, and carbon materials. Efforts are also being made to increase the sensitivity of electrochemical sensors, and the combination of nanomaterials, materials, and biotechnology offers the potential to increase sensitivity in a variety of ways. Recent studies suggest that electrochemical sensors can be a powerful tool in providing comprehensive insights into the targeting and detection of disease-associated biomarkers. Significant advances in nanomaterial and biomolecule approaches for improved sensitivity have resulted in the development of electrochemical biosensors capable of detecting multiple biomarkers in real time in clinically relevant samples. In this review, we have discussed the recent studies on electrochemical sensors for detection of diseases such as diabetes, degenerative diseases, and cancer. Further, we have highlighted new technologies to improve sensitivity using various materials, including DNA, enzymes, nanoparticles, and carbon materials.

Plasmonic nanoparticle amyloid corona for screening Aß oligomeric aggregate-degrading drugs.

The generation of toxic amyloid ß (Aß) oligomers is a central feature of the onset and progression of Alzheimer's disease (AD). Drug discoveries for Aß oligomer degradation have been hampered by the difficulty of Aß oligomer purification and a lack of screening tools. Here, we report a plasmonic nanoparticle amyloid corona (PNAC) for quantifying the efficacy of Aß oligomeric aggregate-degrading drugs. Our strategy is to monitor the drug-induced degradation of oligomeric aggregates by analyzing the colorimetric responses of PNACs. To test our strategy, we use Aß-degrading proteases (protease XIV and MMP-9) and subsequently various small-molecule substances that have shown benefits in the treatment of AD. We demonstrate that this strategy with PNAC can identify effective drugs for eliminating oligomeric aggregates. Thus, this approach presents an appealing opportunity to reduce attrition problems in drug discovery for AD treatment.

Bio-Inspired Electronic Textile Yarn-Based NO2 Sensor Using Amyloid-Graphene Composite.

Graphene-based e-textile gas sensors have received significant attention as wearable electronic devices for human healthcare and environmental monitoring. Theoretically, more the attached graphene on the devices, better is the gas-sensing performance. However, it has been hampered by poor adhesion between graphene and textile platforms. Meanwhile, amyloid nanofibrils are reputed for their ability to improve adhesion between materials, including between graphene and microorganisms. Despite that fact, there has been no attempt to apply amyloid nanofibrils to fabricate graphene-based e-textiles. By biomimicking the adhesion ability of amyloid nanofibrils, herein, we developed a graphene-amyloid nanofibril hybrid e-textile yarn (RGO/amyloid nanofibril/CY) for the detection of NO2. Compared to traditional e-textile yarn, the RGO/amyloid nanofibril/CY showed better performance in response time, sensing efficiency, sensitivity, and selectivity for NO2. Last, we suggested a practical use of RGO/amyloid nanofibril/CY combined with a light-emitting diode as a wearable e-textile gas sensor.

Robust quantum point contact via trench gate modulation.

Quantum point contacts (QPC) are a primary component in mesoscopic physics and have come to serve various purposes in modern quantum devices. However, fabricating a QPC that operates robustly under extreme conditions, such as high bias or magnetic fields, still remains an important challenge. As a solution, we have analyzed the trench-gated QPC (t-QPC) that has a central gate in addition to the split-gate structure used in conventional QPCs (c-QPC). From simulation and modelling, we predicted that the t-QPC has larger and more even subband spacings over a wider range of transmission when compared to the c-QPC. After an experimental verification, the two QPCs were investigated in the quantum Hall regimes as well. At high fields, the maximally available conductance was achievable in the t-QPC due to the local carrier density modulation by the trench gate. Furthermore, the t-QPC presented less anomalies in its DC bias dependence, indicating a possible suppression of impurity effects.

Multiplexed femtomolar detection of Alzheimer's disease biomarkers in biofluids using a reduced graphene oxide field-effect transistor.

Alzheimer's disease (AD) is a neurodegenerative disease that accounts for 70% of all dementia. Early stage diagnosis of AD is essential as there is no certain treatment after the lesion has progressed in the late stage. Nevertheless, there are still limitations of early diagnosis of AD using neuroimaging and psychological memory assessments. Here, we demonstrate ultrasensitive and multiplexed detection of pivotal AD biomarkers (Aß1-42 and t-Tau) in biofluids using a reduced graphene oxide field-effect transistor (gFET). The proposed approach provides a wide logarithmically linear range of detection from 10-1-105 pg mL-1 and a femtomolar-level limit of detection in biofluids (human plasma and artificial cerebrospinal fluid) as well as phosphate-buffered saline (PBS). Furthermore, as these core biomarkers have different surface charges in physiological conditions based on the isoelectric point (pI), we achieved a distinctive output signal for each biomarker. The gFET biosensor platform presented in this paper has great potential and can be used for early diagnosis of AD in clinical practice as well as accurate analysis based on the surface charge of the analytes.

Longitudinal profiling of oligomeric Aß in human nasal discharge reflecting cognitive decline in probable Alzheimer's disease.

Despite clinical evidence indicating a close relationship between olfactory dysfunction and Alzheimer's disease (AD), further investigations are warranted to determine the diagnostic potential of nasal surrogate biomarkers for AD. In this study, we first identified soluble amyloid-ß (Aß), the key biomarker of AD, in patient nasal discharge using proteomic analysis. Then, we profiled the significant differences in Aß oligomers level between patient groups with mild or moderate cognitive decline (n = 39) and an age-matched normal control group (n = 21) by immunoblot analysis and comparing the levels of Aß by a self-standard method with interdigitated microelectrode sensor systems. All subjects received the Mini-Mental State Examination (MMSE), Clinical Dementia Rating (CDR), and the Global Deterioration Scale (GDS) for grouping. We observed higher levels of Aß oligomers in probable AD subjects with lower MMSE, higher CDR, and higher GDS compared to the normal control group. Moreover, mild and moderate subject groups could be distinguished based on the increased composition of two oligomers, 12-mer Aß*56 and 15-mer AßO, respectively. The longitudinal cohort study confirmed that the cognitive decline of mild AD patients with high nasal discharge Aß*56 levels advanced to the moderate stage within three years. Our clinical evidence strongly supports the view that the presence of oligomeric Aß proteins in nasal discharge is a potential surrogate biomarker of AD and an indicator of cognitive decline progression.

Highly Sensitive Micropatterned Interdigitated Electrodes for Enhancing the Concentration Effect Based on Dielectrophoresis.

The concentration effect of dielectrophoresis (DEP) enables detection of biomolecules with high sensitivity. In this study, microstructures were patterned between the interdigitated microelectrodes (IMEs) to increase the concentration effect of DEP. The microstructures increased the electric field gradient ( ∇ | E 2 | ) between the IMEs to approximately 6.61-fold higher than in the bare IMEs with a gap of 10 µm, resulting in a decreased optimal voltage to concentrate amyloid beta 42 (Aß42, from 0.8 Vpp to 0.5 Vpp) and tau-441 (from 0.9 Vpp to 0.6 Vpp) between the IMEs. Due to the concentration effect of DEP, the impedance change in the optimal condition was higher than the values in the reference condition at 2.64-fold in Aß42 detection and at 1.59-fold in tau-441 detection. This concentration effect of DEP was also verified by counting the number of gold (Au) particles which conjugated with the secondary antibody. Finally, an enhanced concentration effect in the patterned IMEs was verified by measuring the impedance change depending on the concentration of Aß42 and tau-441. Our results suggest that microstructures increase the concentration effect of DEP, leading to enhanced sensitivity of the IMEs.

Comparative analyses of plasma amyloid-ß levels in heterogeneous and monomerized states by interdigitated microelectrode sensor system.

Detection of amyloid-ß (Aß) aggregates contributes to the diagnosis of Alzheimer disease (AD). Plasma Aß is deemed a less invasive and more accessible hallmark of AD, as Aß can penetrate blood-brain barriers. However, correlations between biofluidic Aß concentrations and AD progression has been tenuous. Here, we introduce a diagnostic technique that compares the heterogeneous and the monomerized states of Aß in plasma. We used a small molecule, EPPS [4-(2-hydroxyethyl)-1-piperazinepropanesulfonic acid], to dissociate aggregated Aß into monomers to enhance quantification accuracy. Subsequently, Aß levels of EPPS-treated plasma were compared to those of untreated samples to minimize inter- and intraindividual variations. The interdigitated microelectrode sensor system was used to measure plasma Aß levels on a scale of 0.1 pg/ml. The implementation of this self-standard blood test resulted in substantial distinctions between patients with AD and individuals with normal cognition (NC), with selectivity and sensitivity over 90%.

Dielectrophoresis-based filtration effect and detection of amyloid beta in plasma for Alzheimer's disease diagnosis.

The filtration effect improves the impedance change through specific binding of target molecules in plasma, and decreases this change by nonspecific binding of matrix factors in plasma (i.e., matrix effect). A difference in dielectrophoresis (DEP) forces applied to target molecules and matrix factors causes the filtration effect. An optimized DEP force affects target molecules, which remain in the reaction region of an interdigitated microelectrode (IME) sensor. Various matrix factors, which are larger than the target molecules, are influenced by a strong DEP force and are filtered out of the reaction region. To demonstrate the filtration effect, the matrix effect was confirmed in standard plasma and in phosphate-buffered saline, based on the detection of amyloid beta (Aß), an Alzheimer's disease (AD)-associated peptide. The filtration effect was verified using the matrix effect factor (MEF), which was calculated from the impedance change values in different detection environments. In standard plasma, the MEF value decreased by approximately 78.12%, and in buffer with heterogeneous Aß, by approximately 75.43%. Plasma from patients with AD and normal controls (NCs) was analyzed using the value of the impedance change by the filtration effect. The impedance change was enhanced approximately 1.52 ±â€¯0.03-fold in AD plasma, but declined approximately 0.90 ±â€¯0.03-fold in NC plasma. This difference tendency by the filtration effect was the disease evaluation index and used as an important criterion that distinguished between the AD and NC plasma. Plasma-based AD diagnosis may be possible, based on the filtration effect.

Immediate implant placement into infected and noninfected extraction sockets: a pilot study.

OBJECTIVE: This study compared the osseointegration of immediate implants in dogs in infection-free sites and in sites with periradicular lesions which were removed by simulated periradicular surgery. STUDY DESIGN: Periradicular surgeries were performed to remove intentionally induced periradicular lesions, followed by teeth extraction and immediate implant placement with (experimental group 1) or without (experimental group 2) membranes. In the control group, implants were placed at healthy extraction sockets. After 12 weeks, the animals were killed and the results of histomorphometric study were analyzed by Kruskal-Wallis test. RESULTS: Both the control and the experimental implants were clinically acceptable. The control group showed significantly higher bone-implant contact (BIC; 76.03 +/- 7.98%) than the experimental groups 1 (59.55 +/- 14.21%) and 2 (48.62 +/- 20.22%) (P < .05). CONCLUSIONS: Despite the lower BIC of the experimental groups, this pilot study showed the possibility that immediate implant placement might be successful in extraction sockets with periradicular lesions. Further studies with larger sample sizes are required.