Electric-field assisted silicon nanowire field effect transistor for the ultra-low concentration nucleic acid detection.

Ultra-low concentration nucleic acid detection is crucial for disease diagnosis and prognosis. Silicon nanowire field-effect transistors (SiNW FETs) are promising due to their sensitivity, real-time capabilities, and compact design. A critical consideration for FETs is the reaction time required for nucleic acid diffusion to the detection surface, especially at low concentrations.

Intermittent lysis on a single paper-based device to extract exosomal nucleic acid biomarkers from biological samples for downstream analysis.

As the role of exosomes in physiological and pathological processes has been properly perceived, harvesting them and their internal components is critical for subsequent applications. This study is a debut of intermittent lysis, which has been integrated into a simple and easy-to-operate procedure on a single paper-based device to extract exosomal nucleic acid biomarkers for downstream analysis. Exosomes from biological samples were captured by anti-CD63-modified papers before being intermittently lysed by high-temperature, short-time treatment with double-distilled water to release their internal components.

Specific and efficient knockdown of intracellular miRNA using partially neutralized phosphate-methylated DNA oligonucleic acid-loaded mesoporous silica nanoparticles.

Antisense oligonucleotides (ASOs) are molecules used to regulate RNA expression by targeting specific RNA sequences. One specific type of ASO, known as neutralized DNA (nDNA), contains site-specific methyl phosphotriester (MPTE) linkages on the phosphate backbone, changing the negatively charged DNA phosphodiester into a neutralized MPTE with designed locations. While nDNA has previously been employed as a sensitive nucleotide sequencing probe for the PCR, the potential of nDNA in intracellular RNA regulation and gene therapy remains underexplored.

Comparing solution-gate and bottom-gate nanowire field-effect transistors on pH sensing with different salt concentrations and surface modifications.

Field-effect transistors (FETs) have been developed as pH sensors by using various device structures, fabrication technologies, and sensing film materials. Different transistor structures, like extended-gate (EG) FETs, floating-gate FET sensors, and dual-gate (DG) FETs, can enhance the sensor performance. In this article, we report the effects of using solution-gate and bottom-gate FET configurations on pH sensing and investigate the influence of different ionic concentrations of buffers in the measured signals.

Paper-Based Exosomal MicroRNA-21 Detection for Wound Monitoring: A Proof of Concept and Clinical Validation Trial Study.

Emerging evidence has shown that microRNAs play pivotal roles in wound healing. MicroRNA-21 (miR-21) was previously found to upregulate in order to fulfill an anti-inflammation role for wounds. Exosomal miRNAs have been identified and explored as essential markers for diagnostic medicine.

Paper/PMMA hybrid device with a microvalve-controlled design for exosome isolation and analysis.

This study proposes a paper/PMMA hybrid device designed to isolate exosomes and extract exosomal miRNA, followed by quantitative analysis. It aims to provide simplified and convenient sample preparation for potential point-of-care testing (POCT) processes. In contrast to previous work conducted by our research team, which focused on isolating exosomes and exosomal nucleic acids, this study introduces a novel approach by integrating paper and a PMMA mold with a microvalve controlled design.

PLBD: protein-ligand binding database of thermodynamic and kinetic intrinsic parameters.

We introduce a protein-ligand binding database (PLBD) that presents thermodynamic and kinetic data of reversible protein interactions with small molecule compounds. The manually curated binding data are linked to protein-ligand crystal structures, enabling structure-thermodynamics correlations to be determined. The database contains over 5500 binding datasets of 556 sulfonamide compound interactions with the 12 catalytically active human carbonic anhydrase isozymes defined by fluorescent thermal shift assay, isothermal titration calorimetry, inhibition of enzymatic activity and surface plasmon resonance.

Molecular effects of site-specific phosphate-methylated primer on the structure and motions of Taq DNA polymerase.

Polymerase chain reaction (PCR) is a powerful molecular biology assay for gene detection and quantification. Conventional DNA primers for PCR often suffer from poor sensitivity in specific gene detection. Recently, oligonucleotides containing methyl phosphotriester (MPTE-DNA) have been developed with enhanced DNA hybridization and improved gene detection sensitivity.

Ultrasensitive Detection of Interleukin 6 by Using Silicon Nanowire Field-Effect Transistors.

Interleukin 6 (IL-6) has been regarded as a biomarker that can be applied as a predictor for the severity of COVID-19-infected patients. The IL-6 level also correlates well with respiratory dysfunction and mortality risk. In this work, three silanization approaches and two types of biorecognition elements were used on the silicon nanowire field-effect transistors (SiNW-FETs) to investigate and compare the sensing performance on the detection of IL-6.

In-Silico Selection of Aptamer Targeting SARS-CoV-2 Spike Protein.

Aptamers are single-stranded, short DNA or RNA oligonucleotides that can specifically bind to various target molecules. To diagnose the infected cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in time, numerous conventional methods are applied for viral detection via the amplification and quantification of DNA or antibodies specific to antigens on the virus. Herein, we generated a large number of mutated aptamer sequences, derived from a known sequence of receptor-binding domain (RBD)-1C aptamer, specific to the RBD of SARS-CoV-2 spike protein (S protein).

Paper-Based Devices for Capturing Exosomes and Exosomal Nucleic Acids From Biological Samples.

Exosomes, nanovesicles derived from cells, contain a variety of biomolecules that can be considered biomarkers for disease diagnosis, including microRNAs (miRNAs). Given knowledge and demand, inexpensive, robust, and easy-to-use tools that are compatible with downstream nucleic acid detection should be developed to replace traditional methodologies for point-of-care testing (POCT) applications. This study deploys a paper-based extraction kit for exosome and exosomal miRNA analytical system with some quantifying methods to serve as an easy sample preparation for a possible POCT process.

Phosphate-Methylated Oligonucleotides as a Novel Primer for PCR and RT-PCR.

This chapter introduces neutralized DNA (nDNA) as a novel design for the primers of PCR and RT-PCR by methylating phosphate groups of some oligonucleotides in their structures. It starts with an introduction of the nDNA which possesses an electrically chimeric neutral backbone as well as the proposed standards in designing nDNA as a novel primer for PCR and RT-PCR , concluded from various experimental results presented afterward. The primary content comprises empirical data from PCR to compare nDNA and unmodified DNA as primers in terms of ability to distinguish and amplify mismatch templates, activities of polymerase enzymes, melting temperature of double-stranded sequences, and the trials and discussions on various modified positions of the nDNA primers.

Optimizing surface modification of silicon nanowire field-effect transistors by polyethylene glycol for MicroRNA detection.

MicroRNA (miRNA) sensing plays an essential role in the diagnosis of several diseases, especially cancers, for appropriate intervention and treatment. However, quantifying miRNA demands highly sensitive and selective assays which can distinguish analogous sequences with low abundance in bio-samples and determine wide range of concentrations. In this report, we present a novel technique satisfying all those requirements by modifying silicon nanowire field-effect transistors (SiNWFETs) with 2-component mixed self-assembled monolayers (mSAMs) of polyethylene glycol (PEG) at different ratios (silane-PEG-NH:silane-PEG-OH = 1:1, 1:3, and 1:5) and glutaraldehyde to immobilize DNA probes for miRNA-21 detection, a biomarker in several types of cancers.

Switching the Inhibitor-Enzyme Recognition Profile via Chimeric Carbonic Anhydrase XII.

A key part of the optimization of small molecules in pharmaceutical inhibitor development is to vary the molecular design to enhance complementarity of chemical features of the compound with the positioning of amino acids in the active site of a target enzyme. Typically this involves iterations of synthesis, to modify the compound, and biophysical assay, to assess the outcomes. Selective targeting of the anti-cancer carbonic anhydrase isoform XII (CA XII), this process is challenging because the overall fold is very similar across the twelve CA isoforms.

Uncertainty in protein-ligand binding constants: asymmetric confidence intervals versus standard errors.

Equilibrium binding constants (K) between chemical compounds and target proteins or between interacting proteins provide a quantitative understanding of biological interaction mechanisms. Reported uncertainties of measured experimental parameters are critical for decision-making in many scientific areas, e.g.

Combination of Aptamer Amplifier and Antigen-Binding Fragment Probe as a Novel Strategy to Improve Detection Limit of Silicon Nanowire Field-Effect Transistor Immunosensors.

Detecting proteins at low concentrations in high-ionic-strength conditions by silicon nanowire field-effect transistors (SiNWFETs) is severely hindered due to the weakened signal, primarily caused by screening effects. In this study, aptamer as a signal amplifier, which has already been reported by our group, is integrated into SiNWFET immunosensors employing antigen-binding fragments (Fab) as the receptors to improve its detection limit for the first time. The Fab-SiNWFET immunosensors were developed by immobilizing Fab onto Si surfaces modified with either 3-aminopropyltriethoxysilane (APTES) and glutaraldehyde (GA) (Fab/APTES-SiNWFETs), or mixed self-assembled monolayers (mSAMs) of polyethylene glycol (PEG) and GA (Fab/PEG-SiNWFETs), to detect the rabbit IgG at different concentrations in a high-ionic-strength environment (150 mM Bis-Tris Propane) followed by incubation with R18, an aptamer which can specifically target rabbit IgG, for signal enhancement.

Improve sample preparation process for miRNA isolation from the culture cells by using silica fiber membrane.

In clinical applications of miRNAs, the purity and quality of the testing samples are very critical, especially the obtained tissue sample volume is limited. If the extracted miRNAs are contaminated or different in quality before analysis, it will increase the variance of the analysis result and make the medical information judgment incorrect and cannot be portable. Herein, we improved the commercially extraction kit by realizing the fundamental mechanism and hoped to serve finding optimal procedures for increasing the recovery of miRNAs extracted from cultured cells.

Predicting Future Prospects of Aptamers in Field-Effect Transistor Biosensors.

Aptamers, in sensing technology, are famous for their role as receptors in versatile applications due to their high specificity and selectivity to a wide range of targets including proteins, small molecules, oligonucleotides, metal ions, viruses, and cells. The outburst of field-effect transistors provides a label-free detection and ultra-sensitive technique with significantly improved results in terms of detection of substances. However, their combination in this field is challenged by several factors.

Field-Effect Transistor Biosensors for Biomedical Applications: Recent Advances and Future Prospects.

During recent years, field-effect transistor biosensors (Bio-FET) for biomedical applications have experienced a robust development with evolutions in FET characteristics as well as modification of bio-receptor structures. This review initially provides contemplation on this progress by briefly summarizing remarkable studies on two aforementioned aspects. The former includes fabricating unprecedented nanostructures and employing novel materials for FET transducers whereas the latter primarily synthesizes compact molecules as bio-probes (antibody fragments and aptamers).

Signal Enhancement of Silicon Nanowire Field-Effect Transistor Immunosensors by RNA Aptamer.

Silicon nanowire field-effect transistors (SiNW-FETs) have been demonstrated as a highly sensitive platform for label-free detection of a variety of biological and chemical entities. However, detecting signal from immunoassays by nano-FETs is severely hindered by the distribution of different charged groups of targeted entities, their binding orientation, and distances to the surface of the FET. Aptamers have been widely applied as a recognition element for plentiful biosensors because of small molecular sizes and moderate to high specific binding affinity with different types of molecules.

Neutralized chimeric DNA probe for the improvement of GC-rich RNA detection specificity on the nanowire field-effect transistor.

Silicon nanowire (SiNW) field-effect transistors (FETs) is a powerful tool in genetic molecule analysis because of their high sensitivity, short detection time, and label-free detection. In nucleic acid detection, GC-rich nucleic acid sequences form self- and cross-dimers and stem-loop structures, which can easily obtain data containing signals from nonspecific DNA binding. The features of GC-rich nucleic acid sequences cause inaccuracies in nucleic acid detection and hinder the development of precision medicine.

Designed phosphate-methylated oligonucleotides as PCR primers for SNP discrimination.

Polymerase chain reaction (PCR) is a powerful technique for the detection and quantification of nucleic acids and has enormous applications to research in molecular biology. Certain inherited diseases, caused by single nucleotide mutations, however, are difficult to identify by PCR, using DNA primers and probes, in a situation where a false diagnosis may lead to incorrect or delayed treatment. With the aim of enhancing the specificity of PCR, we used novel chemically synthesized oligonucleotides containing site-specific methyl phosphotriester (MPTE) inter-nucleoside linkage(s) as primers and probes.

Repeatability, precision, and accuracy of the enthalpies and Gibbs energies of a protein-ligand binding reaction measured by isothermal titration calorimetry.

In rational drug design, it is important to determine accurately and with high precision the binding constant (the affinity or the change in Gibbs energy, ∆G), the change in enthalpy (ΔH), and the entropy change upon small molecule drug binding to a disease-related target protein. These thermodynamic parameters of the protein-ligand association reaction are usually determined by isothermal titration calorimetry (ITC). Here, the repeatability, precision, and accuracy of the measurement of the affinity and the change in enthalpy upon acetazolamide (AZM) interaction with human carbonic anhydrase II (CA II) are discussed based on the measurements using several ITC instruments.

Synergetic improvements of sensitivity and specificity of nanowire field effect transistor gene chip by designing neutralized DNA as probe.

Neutral DNA analogs as probes for the detection of target oligomers on the biosensors based on the field-effect transistor (FET) configuration feature advantages in the enhancement of sensitivity and signal-to-noise ratio. Herein, we used phosphate-methylated nucleotides to synthesize two partially neutralized chimeric DNA products and a fully neutralized DNA sequence and adopted a regular DNA oligomer as probes on the polycrystalline silicon nanowire (NW) FET devices. The sequences of two neutralized chimeric DNAs close to the 5' end were alternately modified with the phosphate-methylated nucleotides, and all probes were immobilized via their 5' end on the NW surface.

Isothermal titration calorimetry for characterization of recombinant proteins.

Isothermal titration calorimetry is widely used to measure the affinities and enthalpies of interaction between proteins and/or small molecules. The quantitative nature of the technique is especially useful in the characterization of recombinant proteins while determining the fraction of protein capable of binding a specific ligand and thus the protein purity. The revealed thermodynamic information sheds light on the binding mechanism, important for the targeted drug design of the biologics.