Low-Power Wearable Enabled by Extended Gate Field-Effect Transistors to Advance Vigilant Biochemical Sensing.

Biochemical monitoring of sweat through vigilant wearable systems offers new opportunities for improved stress management. A low-power biochemical sensing platform has been developed to perform potentiometric sensing using extended gate field-effect transistors (EGFET) for wearable biochemical monitoring. In vitro validation of the EGFET-enabled electrochemistry was achieved by testing pH and electrolyte concentrations.

An electrochemical biosensor for the detection of tuberculosis specific DNA with CRISPR-Cas12a and redox-probe modified oligonucleotide.

Background: The development of a robust and accurate point-of-care platform for the detection of tuberculosis (TB) biomarkers is important for disease control. In the current study, the detection principle relies on the shredding of PES-modified non-specific ssDNA (Poly T) in the presence of target DNA IS6110, a reliable biomarker for TB diagnosis by the CRISPR-Cas12a mechanism. Cas protein has great potential in the detection of nucleic acids.

Proximity-Unlocked Luminescence by Sequential Enzymatic Reactions from Antibody and Antibody/Aptamer (PULSERAA): A Platform for Detection and Visualization of Virus-Containing Spots.

The SARS-CoV-2 pandemic has challenged more scientists to detect viruses and to visualize virus-containing spots for diagnosis and infection control; however, detection principles of commercially available technologies are not optimal for visualization. Here, a convenient and universal homogeneous detection platform named proximity-unlocked luminescence by sequential enzymatic reactions from antibody and antibody/aptamer (PULSERAA) is developed. This is designed so that the signal appears only when the donor and acceptor are in proximity on the viral surface.

Development of tetravalent antibody-enzyme complexes employing a lactate oxidase and the application to electrochemical immunosensors.

Antibody-enzyme complexes (AECs) are ideal for immunosensing. Although AECs using antibody fragments can be produced by bacterial hosts, their low affinity limits their sensing applications. We have improved the affinity of AECs by combining two antibodies using Catcher/Tag systems; however, it requires multiple antibodies and an enzyme production process.

Levodopa: From Biological Significance to Continuous Monitoring.

A continuous levodopa sensor can improve the quality of life for patients suffering with Parkinson's disease by enhancing levodopa titration and treatment effectiveness; however, its development is currently hindered by the absence of a specific levodopa molecular recognition element and limited insights into how real-time monitoring might affect clinical outcomes. This gap in research contributes to clinician uncertainty regarding the practical value of continuous levodopa monitoring data. This paper examines the current state of levodopa sensing and the inherent limitations in today's methods.

The 2.5 generation enzymatic sensors based on the construction of quasi-direct electron transfer type NAD(P)-Dependent dehydrogenases.

We introduce a versatile method to convert NAD or NADP -dependent dehydrogenases into quasi-direct electron transfer (quasi-DET)-type dehydrogenases, by modifying with a mediator on the enzyme surface toward the development of 2.5 generation enzymatic sensors. In this study, we use β-hydroxybutyrate (BHB) dehydrogenase (BHBDh) from Alcaligenes faecalis (AfBHBDh) as a representative NAD or NADP -dependent dehydrogenase.

Development of closed bipolar electrode based L-lactate sensor employing quasi-direct electron transfer type enzyme with cyclic voltammetry.

Herein, we present a proof-of-concept of an enzyme sensor combining closed bipolar electrode system with quasi-direct electron transfer (DET) type enzyme. The closed bipolar electrode system was tested using cyclic voltammetry, with L-lactate as a model substrate. L-Lactate was detected through measurement of the change in junction potential across the bipolar electrode.

Rapid and Convenient Single-Chain Variable Fragment-Employed Electrochemical C-Reactive Protein Detection System.

Although IgG-free immunosensors are in high demand owing to ethical concerns, the development of convenient immunosensors that alternatively integrate recombinantly produced antibody fragments, such as single-chain variable fragments (scFvs), remains challenging. The low affinity of antibody fragments, unlike IgG, caused by monovalent binding to targets often leads to decreased sensitivity. We improved the affinity owing to the bivalent effect by fabricating a bivalent antibody-enzyme complex (AEC) composed of two scFvs and a single glucose dehydrogenase, and developed a rapid and convenient scFv-employed electrochemical detection system for the C-reactive protein (CRP), which is a homopentameric protein biomarker of systemic inflammation.

Development of Direct Electron Transfer-Type Extended Gate Field Effect Transistor Enzymatic Sensors for Metabolite Detection.

In this work, direct electron transfer (DET)-type extended gate field effect transistor (EGFET) enzymatic sensors were developed by employing DET-type or quasi-DET-type enzymes to detect glucose or lactate in both 100 mM potassium phosphate buffer and artificial sweat. The system employed either a DET-type glucose dehydrogenase or a quasi-DET-type lactate oxidase, the latter of which was a mutant enzyme with suppressed oxidase activity and modified with amine-reactive phenazine ethosulfate. These enzymes were immobilized on the extended gate electrodes.

Development of the BioBattery: A novel enzyme fuel cell using a multicopper oxidase as an anodic enzyme.

This work presents the development of an enzyme fuel cell, termed "BioBattery", that utilizes multicopper oxidases as the anodic enzyme in a non-diffusion limited system. We evaluated various enzyme variants as the anode, including multicopper oxidase from Pyrobaculum aerophilum, laccase from Trametes versicolor, and bilirubin oxidase from Myrothecium verrucaria. Several combinations of cathodes were also examined, focusing on the reduction of oxygen as the primary electron acceptor.

Development of a glycated albumin sensor employing dual aptamer-based extended gate field effect transistors.

Glycated albumin (GA), defined as the percentage of serum albumin glycation, is a mid-term glycemic control marker for diabetes. The concentrations of both glycated human serum albumin (GHSA) and total human serum albumin (HSA) are required to calculate GA. Here, we report the development of a GA sensor employing two albumin aptamers: anti-GHSA aptamer which is specific to GHSA and anti-HSA aptamer which recognizes both glycated and non-glycated HSA.

Exploration and Application of DNA-Binding Proteins to Make a Versatile DNA-Protein Covalent-Linking Patch (D-Pclip): The Case of a Biosensing Element.

DNA-protein complexes are attractive components with broad applications in various research fields, such as DNA aptamer-enzyme complexes as biosensing elements. However, noncovalent DNA-protein complexes often decrease detection sensitivity because they are highly susceptible to environmental conditions. In this study, we developed a versatile DNA-protein covalent-linking patch (D-Pclip) for fabricating covalent and stoichiometric DNA-protein complexes.

Development of an electrochemical impedance spectroscopy immunosensor for insulin monitoring employing pyrroloquinoline quinone as an ingestible redox probe.

Contemporary electrochemical impedance spectroscopy (EIS)-based biosensors face limitations in their applicability for in vivo measurements, primarily due to the necessity of using a redox probe capable of undergoing oxidation and reduction reactions in solution. Although previous investigations have demonstrated the effectiveness of EIS-based biosensors in detecting various target analytes using potassium ferricyanide as a redox probe, its unsuitability for blood or serum measurements, attributed to its inherent toxicity, poses a significant challenge. In response to this challenge, our study adopted a unique approach, focusing on the use of ingestible materials, by exploring naturally occurring substances within the body, with a specific emphasis on pyrroloquinoline quinone (PQQ).

Development of an Anti-Idiotype Aptamer-Based Electrochemical Sensor for a Humanized Therapeutic Antibody Monitoring.

Therapeutic monoclonal antibodies (mAbs) are currently the most effective medicines for a wide range of diseases. Therefore, it is expected that easy and rapid measurement of mAbs will be required to improve their efficacy. Here, we report an anti-idiotype aptamer-based electrochemical sensor for a humanized therapeutic antibody, bevacizumab, based on square wave voltammetry (SWV).

Development of a Versatile Method to Construct Direct Electron Transfer-Type Enzyme Complexes Employing SpyCatcher/SpyTag System.

The electrochemical enzyme sensors based on direct electron transfer (DET)-type oxidoreductase-based enzymes are ideal for continuous and in vivo monitoring. However, the number and types of DET-type oxidoreductases are limited. The aim of this research is the development of a versatile method to create a DET-type oxidoreductase complex based on the SpyCatcher/SpyTag technique by preparing SpyCatcher-fused heme and SpyTag-fused non-DET-type oxidoreductases, and by the in vitro formation of DET-type oxidoreductase complexes.

Microgravity environment grown crystal structure information based engineering of direct electron transfer type glucose dehydrogenase.

The heterotrimeric flavin adenine dinucleotide dependent glucose dehydrogenase is a promising enzyme for direct electron transfer (DET) principle-based glucose sensors within continuous glucose monitoring systems. We elucidate the structure of the subunit interface of this enzyme by preparing heterotrimer complex protein crystals grown under a space microgravity environment. Based on the proposed structure, we introduce inter-subunit disulfide bonds between the small and electron transfer subunits (5 pairs), as well as the catalytic and the electron transfer subunits (9 pairs).

Structure of lactate oxidase from Enterococcus hirae revealed new aspects of active site loop function: Product-inhibition mechanism and oxygen gatekeeper.

l-Lactate oxidase (LOx) is a flavin mononucleotide (FMN)-dependent triose phosphate isomerase (TIM) barrel fold enzyme that catalyzes the oxidation of l-lactate using oxygen as a primary electron acceptor. Although reductive half-reaction mechanism of LOx has been studied by structure-based kinetic studies, oxidative half-reaction and substrate/product-inhibition mechanisms were yet to be elucidated. In this study, the structure and enzymatic properties of wild-type and mutant LOxs from Enterococcus hirae (EhLOx) were investigated.

In Vitro Continuous 3 Months Operation of Direct Electron Transfer Type Open Circuit Potential Based Glucose Sensor: Heralding the Next CGM Sensor.

Background: While continuous glucose monitoring (CGM) systems allow precise and real-time blood glucose control, current electrochemicalbased CGM technologies inherently harbor enzyme instability issues. The direct electron transfer (DET) type open circuit potential (OCP) based enzyme sensing principle can minimize the catalytic turnover of the enzyme reaction, thereby providing longer-term operational stability in future CGM glucose sensors.

Method: DET-type OCP based glucose sensors were constructed using gold disk electrodes with glucose dehydrogenase capable of DET which was immobilized using a self-assembled monolayer (SAM).

Development of an electrochemical impedance spectroscopy based biosensor for detection of ubiquitin C-Terminal hydrolase L1.

Year over year, the incidence of traumatic brain injury (TBI) in the population is dramatically increasing; thus, timely diagnosis is crucial for improving patient outcomes in the clinic. Ubiquitin C-terminal hydrolase L1 (UCH-L1), a blood-based biomarker, has been approved by the FDA as a promising quantitative indicator of mild TBI that arises in blood serum shortly after injury. Current gold standard techniques for its quantitation are time-consuming and require specific laboratory equipment.