Diagnostic Biosensors Research Articles

Microbial biosensors for diagnostics, surveillance and epidemiology: Today's achievements and tomorrow's prospects.

Microbial biosensors hold great promise for engineering high-performance, field-deployable and affordable detection devices for medical and environmental applications. This review explores recent advances in the field, highlighting new sensing strategies and modalities for whole-cell biosensors as well as the remarkable expansion of microbial cell-free systems. We also discuss improvements in robustness that have enhanced the ability of biosensors to withstand the challenging conditions found in biological samples. However, limitations remain in expanding the detection repertoire, particularly for proteins. We anticipate that the AI-powered revolution in protein design will streamline the engineering of custom-made sensing modules and unlock the full potential of microbial biosensors.

Real‐Time Nanoscale Bacterial Detection Utilizing a 1DZnO Optical Nanobiosensor

One‐dimensional zinc oxide nanomaterials (1DZnO) have emerged as promising, cost‐effective nanoplatforms with adjustable properties suitable for electrochemical and optical biosensing applications. In this work, modifications in the inherent photoluminescent response of 1DZnO are harnessed to develop a novel immunosensor tailored for detecting enteropathogenic Escherichia coli. This nanobiosensor demonstrates a modulation in photoluminescence signal, effectively responsive to analyte concentrations ranging from 1 × 102 to 1 × 108 CFU mL−1, with direct visualization of targeted bacterial cells over 1DZnO structures through scanning electron microscopy. The conceptualization of this nanobiosensor is focused on a real‐time contact strategy that can significantly reduce processing and response times for pathogen detection, prospected for emergency scenarios. With this aim, the detection process unfolds in real time, with a mere 5–10 s interaction time, corroborated by the standard polymerase chain reaction approach. This synergistic validation underscores the reliability and precision of the developed biosensor. Notably, the utility of 1DZnO nanoplatforms extends beyond the realm of enteropathogenic E. coli, as the biosensing performance exhibited here holds promise for analogous applications involving other medically pertinent pathogens. This study paves the way for the broader implementation of 1DZnO‐based biosensors in medical diagnostics, offering rapid, sensitive, and real‐time detection capabilities.

Allosteric stem-loop multicomponent DNAzyme: A versatile stimuli-responsive switch for nanotweezer operations and multifunctional biosensing

DNAzyme, possessing the capabilities of both encoding information and catalyzing chemical reactions, holds significant value in advancing the widespread application of dynamic nanotechnology. Particularly, multicomponent DNAzyme (MNAzyme) has attracted considerable attention due to its remarkable versatility and practicality. However, the inevitable signal leakage of MNAzyme constrains the applications in complex nucleic acid reaction networks. Herein, we present an allosteric stem-loop multicomponent DNAzyme (ASLM) that incorporates a blocking domain to mitigate leakage resulting from secondary structures of MNAzyme while simultaneously stabilizing binding affinity during complex formation. As the sequence of blocking domain serves as an extension independent of the functional region in MNAzyme, this modulation structure can be flexibly assembled with conformation-switchable nanotweezer to form a time-responsive molecular device, enabling dynamic and dissipative operations through nucleic acid and RNase H. More importantly, the ASLM structure can be directly incorporated into DNA catalytic circuits for detecting multiple biomolecules (nucleic acid, protease, and small molecule). With its scalability and versatility, this effector-activated allosteric regulator will serve as a universal stimuli-responsive switch in complex and hierarchical DNA networks, expanding the scope of DNA nanotechnology in molecular device construction and diagnostic biosensing.

Biosensing platforms for DNA diagnostics based on CRISPR/Cas nucleases: towards the detection of nucleic acids at the level of single molecules in non-laboratory settings.

The use of CRISPR/Cas nucleases for the development of DNA diagnostic systems in out-of-laboratory conditions (point-of-need testing, PONT) has demonstrated rapid growth in the last few years, starting with the appearance in 2017-2018 of the first diagnostic platforms known as DETECTR and SHERLOCK. The platforms are based on a combination of methods of nucleic acid isothermal amplification with selective CRISPR/Cas detection of target amplicons. This significantly improves the sensitivity and specificity of PONT, making them comparable with or even superior to the sensitivity and specificity of polymerase chain reaction, considered as the "gold standard" of DNA diagnostics. The review considers modern approaches to the coupling of CRISPR/Cas detection using Cas9, Cas12a, Cas12b, Cas13a, Cas14, and Cas3 nucleases to various methods of nucleic acid isothermal amplification, with an emphasis on works in which sensitivity at the level of single molecules (attomolar and subattomolar concentrations of the target) is achieved. The properties of CRISPR/Cas nucleases used for targeted DNA diagnostics and the features of methods of nucleic acid isothermal amplification are briefly considered in the context of the development of diagnostic biosensing platforms. Special attention is paid to the most promising directions for the development of DNA diagnostics using CRISPR/Cas nuclease.

(Invited) MoS2 Photodetectors for Near-Infrared Biosensing Applications

Device researchers have been actively developing novel diagnostic biosensors using metallic nanoparticle-based plasmonic immunoassays.[1] In these biosensing devices, photodetectors with high photoresponsivity and low internal noise levels are integrated to detect the marginal optical signals change induced by biomarker binding events (i.e., Antigen-antibody binding). Recently, atomically thin layers of molybdenum disulfide (MoS2) have been integrated with plasmonic components to enable fast and sensitive colorimetric monitoring of disease-related biomarkers.[2] However, such biosensors are mostly operated in the ultraviolet/visible range, which needs a purification step to separate out a variety of unwanted biomaterials that absorb visible light and takes several hours of preparation steps. Thus, it would be amenable to develop biosensors for biomolecular detections or immunoassays in the whole blood (WB) analytes without any sample preparations.[3] To address the issues mentioned above, recent studies have demonstrated the engineered gold nanoparticles (AuNPs) that have localized surface plasmonic resonance (LSPR) shift around near-infrared (NIR) wavelength region. However, a high-sensitivity photodetector under NIR region is still required to detect marginal signal changes due to the target biomarker binding events. Meanwhile, the MoS2 material has been reported to exhibits superior photo-response characteristics.[4] MoS2 is a transition metal dichalcogenide material of which single- and multi-layer films exhibit an efficient electron-hole pair generation rate under photoexcitation and therefore high photo absorption as compared to silicon. Therefore, a systematical study on MoS2 photoconductors to optimize the optoelectronic properties under near-infrared (NIR) (λ = 650 nm) operation can enable zero-preparation WB immunoassays combined with specially engineered AuNPs.In this work, we study the photo-response properties (i.e., Photoresponsivity spectrum) of in-plane MoS2 photodetectors as the function of their geometric dimensions and fabrication conditions. Recent study shows that an annealing temperature after exfoliation can etch the upper layers of MoS2 flakes and clean stains on the device and therefore improve device performances (e.g., mobility). [5] This work enables the NIR operation capabilities of plasmonic colorimetric biosensing by introducing the optimized MoS2 photodetector fabrication practice. This approach reduces assay preparation times and mitigate background interference even with WB analytes and thereby enable the WB point-of-care immunoassays.

Electrochemical biosensors for clinical detection of bacterial pathogens: advances, applications, and challenges.

Bacterial pathogens are responsible for a variety of human diseases, necessitating their prompt detection for effective diagnosis and treatment of infectious diseases. Over recent years, electrochemical methods have gained significant attention owing to their exceptional sensitivity and rapidity. This review outlines the current landscape of electrochemical biosensors employed in clinical diagnostics for the detection of bacterial pathogens. We categorize these biosensors into four types: amperometry, potentiometry, electrochemical impedance spectroscopy, and conductometry, targeting various bacterial components, including toxins, virulence factors, metabolic activity, and events related to bacterial adhesion and invasion. We discuss the merits and challenges associated with electrochemical methods, underscoring their rapid response, high sensitivity, and specificity, while acknowledging the necessity for skilled operators and potential interference from biological and environmental factors. Furthermore, we examine future prospects and potential applications of electrochemical biosensors in clinical diagnostics. While electrochemical biosensors offer a promising avenue for detecting bacterial pathogens, further research in optimizing the robustness and surmounting the challenges hindering their seamless integration into clinical practice is imperative.

Opto-electrochemical transducers for molecular sensing utilizing nanozyme-based sensing technologies

As a potential substitute for naturally occurring enzymes, nanozymes (nanomaterials with enzyme-mimicking catalytic activity) have emerged in recent years. Natural enzymes have a number of intrinsic shortcomings that could be addressed by nanozymes, including their expensive price, poor stability, storage complexities, and reliance on particular working conditions (e.g., restricted substrate, pH, and temperature ranges). Therefore, the creation and deployment of sophisticated nanozymes has the potential to drastically alter the state of biosensor applications and the life sciences as they exist today. Biosensors have been utilized for quite some time as a streamlined method to quantitatively detect biomarkers (analytes) of interest. Therefore, the utilization of nanozymes in such biosensors assumes significant importance in the pursuit of cost-effective and dependable biosensors for therapeutic monitoring, disease prognosis, and diagnostics. Nanozymes have been investigated for a wide range of biosensing applications throughout the last few years. However, only few of such initiatives have produced platforms and devices that may be used for point-of-care and on-site monitoring. Hence, by providing a thorough overview of current developments in the classification of common nanozymes, this review seeks to guide and support the future development of biosensor technologies based on nanozymes, with an emphasis on their applications in the biosensing sector. We envision that this review will promote coordinated efforts across disciplines and sectors, helping to bridge the gap that keeps nanozyme-based biosensor technology confined to lab settings rather than being used in real-world situations.

Electrochemical aptasensor based on RGO/PPy/Au*NPs for diagnostic biosensing of SARS-CoV-2 in clinical samples: Experimental and molecular dynamics simulations approaches

A highly sensitive and selective E-DNA biosensor was fabricated on a gold substrate using a reduced graphene oxide/polypyrrole/gold nanoparticle/oligonucleotide (RGO/PPy/Au*NPs/Apt) nanocomposite electrode to detect the nucleocapsid protein of SARS-CoV-2 in the patient blood plasma. The modified electrode was characterized by physicochemical techniques such as Fourier transform-infra red (FT-IR), Raman spectroscopy, X-ray diffraction (XRD), energy-dispersive X-ray (EDX) spectroscopy, X-ray photoelectron spectroscopy (XPS) analyses, Brunauer-Emmett-Teller (BET), and Barrett-Joyner-Halenda (BJH) analyses. Moreover, electrochemical analyses were employed to study the electrochemical performance of the electrodes including cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and square wave voltammetry (SWV). Computational investigations of N protein bindings such as hydrogen bonding, Van der Waals binding, and Gibbs binding energies of the N-protein, and aptamer were studied by Molecular Dynamics Simulations (MDS). The unique synergistic effect of RGO, PPy, and the well-known effect of Au nanoparticles makes the DNA probe immobilized on the gold electrode surface. After optimizing the systems, the E-DNA biosensor exhibited a fast SWV response, higher sensitivity (33.77 μA.nM−1.cm−2) and selectivity, high efficiency, good storage stability, and acceptable repeatability for monitoring DNA. The results of real samples based on SWV indicated the correct functioning of the aptasensor in the presence of the SARS-CoV-2 virus. The limit of detection was 3.16×10−17 M and the limit of quantitation was 1.42×10−16 M. The MDS results indicated the stable dynamic folding of the aptamer for beneficial binding. The results indicated that the RGO/PPy/Au*NPs/Apt biosensor is promising for detecting of SARS-CoV-2 virus.

A Clinically Feasible Diagnostic Electrochemical Micronano Motors Biosensor Built on Miniature Swimmer for Multiplex Detection and Grading of Breast Cancer Biomarkers.

Estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor-2 (HER2), and Ki67 are four crucial biomarkers used in the clinical diagnosis of breast cancer. Accurate detection of these biomarkers is essential for an effective diagnosis and treatment. MOF-based micronano motors (MOFtors) are promising for various applications, including environmental remediation, targeted nanosurgery, and biomarker detection. This paper presents a clinically feasible diagnostic electrochemical micronano motor biosensor, built on a miniature swimmer, for the multiplex detection and grading of breast cancer biomarkers. We designed a biosensor, named MOFtor-MSEM, incorporating aptamers and antibodies functionalized on SiO2@Co-Fe-MOF, which acts as a miniature swimmer in solution. The SiO2@Co-Fe-MOF serves as the body, while complementary double-chain-linked antibodies function as paddles. In a homogeneous solution, when a positive voltage is applied to the working electrode, the electrostatic interaction between the neutral SiO2@Co-Fe-MOF and the negatively charged complementary double-linked antibody causes the antibody to move toward the electrode and then regress due to water resistance. This back-and-forth motion propels the miniature swimmer, enabling it to move the target analyte through the solution. The sensor features an automatic "sample-amplifying signal-output" process, achieving simultaneous signal amplification and output of four electrochemical signals on a single nanomaterial, a significant challenge in electrochemical sensing. The biosensor boasts a short detection time of 40 min, compared to approximately 1 week for current clinical tissue testing. Additionally, the bioplatform selectively detects HER2, ER, Ki67, and PR in the range of 0-1500 pg/mL, with detection limits of 0.01420, 0.03201, 0.01430, and 0.01229 pg/mL, respectively.

Exploration of new ways for CRISPR/Cas12a activation: DNA hairpins without PAM and toehold and single strands containing DNA and RNA bases

The CRISPR/Cas12a system is emerging as a promising candidate for next-generation diagnostic biosensing platforms, with the discovery of new activation modes greatly expanding its applications. Here, we have identified two novel CRISPR/Cas12a system activation modes: PAM- and toehold-free DNA hairpins, and DNA-RNA hybrid strands. Utilizing a well-established real-time fluorescence method, we have demonstrated a strong correlation between DNA hairpin structures and Cas12a activation. Compared with previously reported activation modes involving single-stranded DNA and PAM-contained double-stranded DNA, the DNA hairpin activation way exhibits similar specificity and generality. Moreover, our findings indicate that increasing the number of RNA bases in DNA-RNA hybrid strands can decelerate the kinetics of Cas12a-triggered trans-cleavage of reporter probes. These newly discovered CRISPR/Cas12a activation ways hold significant potential for the development of high-performance biosensing strategies.

Filtered Saliva for Rapid and Accurate Analyte Detection for POC Diagnostics.

Saliva has shown considerable promise as a diagnostic medium for point-of-care (POC) and over-the-counter (OTC) diagnostic devices due to the non-invasive nature of its collection. However, a significant limitation of saliva-based detection is undesirable interference in a sensor's readout caused by interfering components in saliva. In this study, we develop standardized sample treatment procedures to eliminate bubbles and interfering molecules while preserving the sample's target molecules such as spike (S) protein and glucose. We then test the compatibility of the pretreatment system with our previously designed SARS-CoV-2 and glucose diagnostic biosensing systems for detecting S protein and glucose in subject saliva. Ultimately, the effectiveness of each filter in enhancing biomarker sensitivity is assessed. The results show that a 20 mg nylon wool (NW) filter shows an 80% change in viscosity reduction with only a 6% reduction in protein content, making it an appropriate filter for the salivary S protein diagnostic system. Meanwhile, a 30 mg cotton wool (CW) filter is identified as the optimal choice for salivary glucose detection, achieving a 90% change in viscosity reduction and a 60.7% reduction in protein content with a minimal 4.3% reduction in glucose content. The NW pretreatment filtration significantly improves the limit of detection (LOD) for salivary S protein detection by five times (from 0.5 nM to 0.1 nM) and it reduces the relative standard deviation (RSD) two times compared to unfiltered saliva. Conversely, the CW filter used for salivary glucose detection demonstrated improved linearity with an R2 of 0.99 and a sensitivity of 36.6 μA/mM·cm2, over twice as high as unfiltered saliva. This unique filtration process can be extended to any POC diagnostic system and optimized for any biomarker detection, making electrochemical POC diagnostics more viable in the current market.

Green biosynthesis of silver nanoparticles from aqueous Zingiber montanum flower extract for enhanced application in medical diagnostic biosensors

Green biosynthesis of silver nanoparticles from aqueous Zingiber montanum flower extract for enhanced application in medical diagnostic biosensors

Development of smart core–shell nanoparticles-based sensors for diagnostics of salivary alpha-amylase in biomedical and forensics

Abstract Smart biocompatible materials that respond to a variety of external stimuli have a lot of potential in the creation of low-cost diagnostic biosensors. The present work describes the creation of core–shell nanoparticles as a biosensor for smart enzyme detection of salivary alpha-amylase (sAA). A chitosan-tripolyphosphate core was generated via ionic gelation and was coated with a starch–iodine shell to create biocompatible core–shell nanoparticles. The starch–iodine shell was ruptured in the presence of certain amounts of amylase, exposing the core. This application explains a noticeable color change from blue to white that can be used to identify sAA at the point of care. Synthesized nanoparticles were examined for scanning electron microscopy analysis and energy-dispersive X-ray (EDX). An EDX report reveals that the nanoparticles have higher carbon content at 55% followed by an oxygen atom of 35%. Fourier-transform infrared spectroscopic analysis revealed that the core–shell nanoparticles have carbonyl (C═O) functional groups present. A confirmatory test of amylase reaction on nanoparticle-impregnated paper turns blue to white indicating that the nanoparticle reacts with amylase as an indicator. This paper-based method can be used in future applications in forensic and medical applications.

The importance, benefits, and future of nanobiosensors for infectious diseases.

Infectious diseases, caused by pathogenic microorganisms such as bacteria, viruses, parasites, or fungi, are crucial for efficient disease management, reducing morbidity and mortality rates and controlling disease spread. Traditional laboratory-based diagnostic methods face challenges such as high costs, time consumption, and a lack of trained personnel in resource-poor settings. Diagnostic biosensors have gained momentum as a potential solution, offering advantages such as low cost, high sensitivity, ease of use, and portability. Nanobiosensors are a promising tool for detecting and diagnosing infectious diseases such as coronavirus disease, human immunodeficiency virus, and hepatitis. These sensors use nanostructured carbon nanotubes, graphene, and nanoparticles to detect specific biomarkers or pathogens. They operate through mechanisms like the lateral flow test platform, where a sample containing the biomarker or pathogen is applied to a test strip. If present, the sample binds to specific recognition probes on the strip, indicating a positive result. This binding event is visualized through a colored line. This review discusses the importance, benefits, and potential of nanobiosensors in detecting infectious diseases.

Advanced Nanomaterial-Based Biosensors for N-Terminal Pro-Brain Natriuretic Peptide Biomarker Detection: Progress and Future Challenges in Cardiovascular Disease Diagnostics.

Cardiovascular diseases (CVDs) represent a significant challenge in global health, demanding advancements in diagnostic modalities. This review delineates the progressive and restrictive facets of nanomaterial-based biosensors in the context of detecting N-terminal pro-B-type natriuretic peptide (NT-proBNP), an indispensable biomarker for CVD prognosis. It scrutinizes the escalation in diagnostic sensitivity and specificity attributable to the incorporation of novel nanomaterials such as graphene derivatives, quantum dots, and metallic nanoparticles, and how these enhancements contribute to reducing detection thresholds and augmenting diagnostic fidelity in heart failure (HF). Despite these technological strides, the review articulates pivotal challenges impeding the clinical translation of these biosensors, including the attainment of clinical-grade sensitivity, the substantial costs associated with synthesizing and functionalizing nanomaterials, and their pragmatic deployment across varied healthcare settings. The necessity for intensified research into the synthesis and functionalization of nanomaterials, strategies to economize production, and amelioration of biosensor durability and ease of use is accentuated. Regulatory hurdles in clinical integration are also contemplated. In summation, the review accentuates the transformative potential of nanomaterial-based biosensors in HF diagnostics and emphasizes critical avenues of research requisite to surmount current impediments and harness the full spectrum of these avant-garde diagnostic instruments.

Development of Biosensor for early Detection of COVID-19

Infectious disease such as COVID-19 is one of the major concerns in Malaysia as it becomes the second killing disease and causes huge number of death and spread to other regions of the world if left unchecked. In developed countries, infectious diseases are often preventable, but lack of medical devices in detecting it makes the death cases increase. The growth of different COVID-19 mutation has given so much challenges in detecting, preventing and curing. This gives motivation to researchers in order to solve this global problem by creating and advancing the detection tools and methods. Time, equipment availability, and the biological nature of COVID-19 influence the selection of appropriate detection techniques. This paper summarizes the comprehensive review on the type of diagnostic tests and biosensors available in detecting COVID-19 disease.

The clinical potential of l-oligonucleotides: challenges and opportunities.

Chemically modified nucleotides are central to the development of biostable research tools and oligonucleotide therapeutics. In this context, l-oligonucleotides, the synthetic enantiomer of native d-nucleic acids, hold great promise. As enantiomers, l-oligonucleotides share the same physical and chemical properties as their native counterparts, yet their inverted l-(deoxy)ribose sugars afford them orthogonality towards the stereospecific environment of biology. Notably, l-oligonucleotides are highly resistant to degradation by cellular nucleases, providing them with superior biostability. As a result, l-oligonucleotides are being increasingly utilized for the development of diverse biomedical technologies, including molecular imaging tools, diagnostic biosensors, and aptamer-based therapeutics. Herein, we present recent such examples that highlight the clinical potential of l-oligonucleotides. Additionally, we provide our perspective on the remaining challenges and practical considerations currently associated with the use of l-oligonucleotides and explore potential solutions that will lead to the broader adoption of l-oligonucleotides in clinical applications.

SERS-based microdevices for use as in vitro diagnostic biosensors.

Advances in surface-enhanced Raman scattering (SERS) detection have helped to overcome the limitations of traditional in vitro diagnostic methods, such as fluorescence and chemiluminescence, owing to its high sensitivity and multiplex detection capability. However, for the implementation of SERS detection technology in disease diagnosis, a SERS-based assay platform capable of analyzing clinical samples is essential. Moreover, infectious diseases like COVID-19 require the development of point-of-care (POC) diagnostic technologies that can rapidly and accurately determine infection status. As an effective assay platform, SERS-based bioassays utilize SERS nanotags labeled with protein or DNA receptors on Au or Ag nanoparticles, serving as highly sensitive optical probes. Additionally, a microdevice is necessary as an interface between the target biomolecules and SERS nanotags. This review aims to introduce various microdevices developed for SERS detection, available for POC diagnostics, including LFA strips, microfluidic chips, and microarray chips. Furthermore, the article presents research findings reported in the last 20 years for the SERS-based bioassay of various diseases, such as cancer, cardiovascular diseases, and infectious diseases. Finally, the prospects of SERS bioassays are discussed concerning the integration of SERS-based microdevices and portable Raman readers into POC systems, along with the utilization of artificial intelligence technology.

Application of Antimicrobial Peptides as Diagnostic Biosensors.

The COVID-19 pandemic has shown how emerging infectious diseases could quickly affect the global health and economy. New pathogens with pandemic potential are also expected to appear soon. Moreover, the large use of antibiotics has led to the development of different so-called "superbugs" capable of escaping all of the current antibiotics. In this context, the early and cost-effective detection of pathogens is crucial to avoid the spreading of new pathogens. Here, we present molecular sensors for the recognition of a broad panel of different bacterial species. The detection is based on the use of bacteria-binding peptides (BBPs) in combination with horseradish peroxidase (HRP). We developed a reliable ELISA-like assay that permits us to study the affinity of different BBPs toward some of the most important bacterial pathogens.

Recent advances on portable photoelectrochemical biosensors for diagnostics

AbstractEfficient and accurate diagnostic methods are important for human health monitoring and prevention of major diseases, so the establishment of efficient and convenient testing methods for clinical diagnosis is urgent. Photoelectrochemical (PEC) biosensors have attracted much attention because of their high sensitivity, low detection background, and especially the advantages of combining photoexcitation and electrochemical detection. With the rapid development of integrated circuit technology many wireless transmissions of highly aggregated portable micro devices are manufactured, its small size, easy to carry, and the ability to visualize the results of the test is shown to be the potential for the application of such devices. This review aims to cover recent advances in PEC biosensing within the field of portable detection, including the principles of PEC biosensors, common photovoltaic materials, and scalable fabrication techniques for portable PEC biosensing. Future prospects in the field are also discussed.