Point-of-care dengue detection: polydopamine-modified electrode for rapid NS1 protein testing for clinical samples.

Early diagnosis of dengue infection by detecting the dengue virus non-structural protein 1 (DENV-NS1) is important to the patients to initiate speedy treatment. Enzyme-linked immunosorbent assay (ELISA)-based NS1 detection and RT-PCR are time-consuming and too complex to be employed in remote areas of dengue-endemic countries. Meanwhile, those of NS1 rapid test by lateral flow assay suffer from low detection limit. Electrochemical-based biosensors using screen-printed gold electrodes (SPGEs) have become a reliable detection method to convey both ELISA's high sensitivity and rapid test portability. In this research, we developed an electrochemical biosensor for DENV-NS1 detection by employing polydopamine (PDA)-modified SPGE. The electrodeposition of PDA on the surface of SPGE serves as a bioconjugation avenue for anti-NS1 antibody through a simple and low-cost immobilization procedure. The biosensor performance was evaluated to detect DENV-NS1 protein in PBS and human serum through a differential pulse voltammetric (DPV) technique. The developed sensing platform displayed a low limit of detection (LOD) of 1.63 pg mL-1 and a wide linear range of 10 pg mL-1 to 1 ng mL-1 (R2 ∼ 0.969). The sensing platform also detected DEV-NS1 from four different serotypes in the clinical samples collected from dengue patients in India and Indonesia, with acceptable sensitivity, specificity, and accuracy values of 90.00%, 80.95%, and 87.65%, respectively. This result showcased the facile and versatile method of PDA coating onto the surface of screen-printed gold electrodes for a miniaturized point-of-care (PoC) detection device.

Development of disposable electrode for the detection of mosquito-borne viruses.

Development of disposable, rapid, and convenient biosensor with high sensitivity and reliability is the most desired method of viral disease prevention. To achieve this goal, in this work, a practical impedimetric biosensor has been implemented into a disposable electrode on a screen-printed carbon electrode (SPCE) for the detection of two mosquito-borne viruses. The biosensor fabrication has step-wisely carried out on the disposable electrode surface at room temperature: starting from conductive film formation, physical binding of the gold nanoparticles (AuNPs)-polyaniline (PAni) into the conductive film, and biofunctionalization. To get the maximum efficiency of the antibody, biotinylated antibody has been conjugated on the surface of AuNP-PAni/PAni-SPCE via the streptavidin-biotin conjugation method which is a critical factor for the high sensitivity. Using the antibody-antigen interaction, this disposable electrode has designed to detect mosquito-borne infectious viruses, Chikungunya virus (CHIKV), and Zika virus (ZIKV) separately in a wide linear range of 100 fg mL-1 to 1 ng mL-1 with a low detection limit of 1.33 and 12.31 fg mL-1 , respectively.

CdSe-Co3O4@TiO2 nanoflower-based photoelectrochemical platform probing visible light-driven virus detection.

The design and construction of a visible light-driven photoelectrochemical (PEC) device is described based on a CdSe-Co3O4@TiO2 nanoflower (NF). Moreover, an application to the ultrasensitive detection of viruses, such as hepatitis E virus (HEV), HEV-like particles (HEV-LPs), and SARS-CoV-2 spike protein in complicated lysate solution, is demonstrated. The photocurrent response output of a PEC device based on CdSe-Co3O4@TiO2 is enhanced compared with the individual components, TiO2 and CdSe-Co3O4. This can be attributed to the CdSe quantum dot (QD) sensitization effect and strong visible light absorption to improve overall system stability. A robust oxygen-evolving catalyst (Co3O4) coupled at the hole-trapping site (CdSe) extends the interfacial carrier lifetime, and the energy conversion efficiency was improved. The effective hybridization between the antibody and virus resulted in a linear relationship between the change in photocurrent density and the HEV-LP concentration ranging from 10 fg mL-1 to 10 ng mL-1, with a detection limit of 3.5 fg mL-1. This CdSe-Co3O4@TiO2-based PEC device achieved considerable sensitivity, good specificity, and acceptable stability and demonstrated a significant ability to develop an upgraded device with affordable and portable biosensing capabilities.

Improvement of Modular Protein Display Efficiency in SpyTag-Implemented Norovirus-like Particles.

Genetic fusion and chemical conjugation are the most common approaches for displaying a foreign protein on the surface of virus-like particles (VLPs); however, these methods may negatively affect the formation and stability of VLPs. Here, we aimed to develop a modular display platform for protein decoration on norovirus-like particles (NoV-LPs) by combining the NoV-LP scaffold with the SpyTag/SpyCatcher bioconjugation system, as the NoV-LP is an attractive protein nanoparticle to carry foreign proteins for various applications. The SpyTagged-NoV-LPs were prepared by introducing SpyTag peptide into the C-terminus of the norovirus VP1 protein. To increase surface exposure of the SpyTag peptide on the NoV-LPs, two or three repeated extension linkers (EAAAK) were inserted between the SpyTag peptide and VP1 protein. Fluorescence proteins, EGFP and mCherry, were fused to SpyCatcher and employed as SpyTag conjugation partners. These VP1-SpyTag variants and SpyCatcher-fused EGFP and mCherry were separately expressed in silkworm fat bodies and purified. This study reveals that adding an extension linker did not disrupt the VLP formation; instead, it increased the particle size by 4-6 nm. The conjugation efficiency of the VP1-SpyTag variants with the extended linker improved from ∼15-35 to ∼50-63% based on the densitometric analysis, while it was up to 77% based on an optical quantification of EGFP and mCherry. Results indicate that the linker causes the SpyTag peptides to be positioned further away from the C-termini of VP1 and potentially increases the exposure of the SpyTag to the outer surface of the NoV-LPs, allowing more SpyTag/SpyCatcher complex formation on the VLP surface. Our study provides a strategy for enhancing the conjugation efficiency of NoV-LP and demonstrates the platform's utility for developing vaccines or functional nanoparticles.

Pt-embodiment ZIF-67-derived nanocage as enhanced immunoassay for infectious virus detection.

A facile and general strategy has been employed to develop highly-active nanozyme for immunoassay purposes. The hollow nanostructure of the Co3O4 nanocages (NCs) was anchoring the platinum nanoparticles (PtNPs) enclosed by the exposed oxides framework nd formed PtNPs@Co3O4 NCs. The embodiment of PtNPs was considered an ideal hybrid nanozyme that efficiently catalyzed the oxidation of the substrate molecules with enhanced activity. The PtNPs@Co3O4 NCs were revisited and repurposed on showing its nanozyme's activity with optimization done for the immunoassay platform. The embodiment of 32.44% Pt in the hollow nanostructures demonstrated the highest signal-to-noise responses in the immunoassay. In addition, the stepwise analysis highlighted the enhancement factor of the nanocages' catalytic mechanism. Based on their catalytic activity, these nanocages have been demonstrated to enable sub-femtogram level biosensing of norovirus-like particles (NoV-LPs) with highly selective signals in the capture-detect immunoassay format. The detection limit of the prepared immunoassay achieved 33.52 viral NoV copies/mL of the detection limit, which is 321-folds lower magnitude of the commercial ELISA. This nanocage's enhanced synergic catalytic properties could have great potential applications, including catalysis, biological labeling, and bioassays.

Advancement of dengue virus NS1 protein detection by 3D-nanoassembly complex gold nanoparticles utilizing competitive sandwich aptamer on disposable electrode.

Signal amplification have been centralized in developing the highly reliable biosensor for analyte detection with a narrow detection window. We proposed an aptasensor to provide a highly sensitive early-stage diagnostic platform of dengue virus NS1 protein (DENV-NS1) by dual-approach - colorimetric and electrochemical detection. This work utilized two different aptamers specific to DENV-NS1: One conjugated to gold nanoparticles (AuNPs), forming AuNPs-Apt1 and its complementary sequence aptamer, forming AuNPs-Apt2. The unbound Apt1 of AuNPs-Apt1 by DENV-NS1 were to hybridize to AuNPs-Apt2 and induced a 3D-nanoassembled formation, resulting in DENV-NS1 concentration-dependent plasmonic color change. Occurrence of the hybridization of Apt1 and Apt2, the 3D-assembled hybridized aptamers of AuNPs was incubated with methylene blue (MB) solution, which intercalated a high number of MB molecules within the duplex structure of aptamers, and the complex was captured on the Apt2-conjugated disposable gold electrode (DGE). The developed aptamer-based biosensor showed high sensitivity with colorimetric response down to 1.28 pg/mL and electrochemical approach down to 30 fg/mL of DENV-NS1 with good selectivity. This work showcases an advanced utilization of aptamer and its complementary anti-sense aptamer in signal amplification and nanocarrier for biosensing.

Nanosphere Structures Using Various Materials: A Strategy for Signal Amplification for Virus Sensing.

Nanomaterials have been explored in the sensing research field in the last decades. Mainly, 3D nanomaterials have played a vital role in advancing biomedical applications, and less attention was given to their application in the field of biosensors for pathogenic virus detection. The versatility and tunability of a wide range of nanomaterials contributed to the development of a rapid, portable biosensor platform. In this review, we discuss 3D nanospheres, one of the classes of nanostructured materials with a homogeneous and dense matrix wherein a guest substance is carried within the matrix or on its surface. This review is segmented based on the type of nanosphere and their elaborative application in various sensing techniques. We emphasize the concept of signal amplification strategies using different nanosphere structures constructed from a polymer, carbon, silica, and metal-organic framework (MOF) for rendering high-level sensitivity of virus detection. We also briefly elaborate on some challenges related to the further development of nanosphere-based biosensors, including the toxicity issue of the used nanomaterial and the commercialization hurdle.

Self-Assembled Chromogenic Polymeric Nanoparticle-Laden Nanocarrier as a Signal Carrier for Derivative Binary Responsive Virus Detection.

In the current biosensor, the signal generation is limited to single virus detection in the reaction chamber. An adaptive strategy is required to enable the recognition of multiple viruses for diagnostics and surveillance. In this work, a nanocarrier is deployed to bring specific signal amplification into the biosensor, depending on the target viruses. The nanocarrier is designed using pH-sensitive polymeric nanoparticle-laden nanocarriers (PNLNs) prepared by sequential nanoprecipitation. The nanoprecipitation of two chromogens, phenolphthalein (PP) and thymolphthalein (TP), is investigated in three different solvent systems in which PNLNs demonstrate a high loading of the chromogen up to 59.75% in dimethylformamide (DMF)/dimethyl sulfoxide (DMSO)/ethanol attributing to the coprecipitation degree of the chromogens and the polymer. The PP-encapsulated PNLNs (PP@PNLNs) and TP-encapsulated PNLNs (TP@PNLNs) are conjugated to antibodies specific to target viruses, influenza virus A subtype H1N1 (IV/A/H1N1) and H3N2 (IV/A/H3N2), respectively. After the addition of anti-IV/A antibody-conjugated magnetic nanoparticles (MNPs) and magnetic separation, the enriched PNLNs/virus/MNPs sandwich structure is treated in an alkaline solution. It demonstrates a synergy reaction in which the degradation of the polymeric boundary and the pH-induced colorimetric development of the chromogen occurred. The derivative binary biosensor shows feasible detection on IV/A with excellent specificities of PP@PNLNs on IV/A/H1N1 and TP@PNLNs on IV/A/H3N2 with LODs of 27.56 and 28.38 fg mL-1, respectively. It intrigues the distinguished analytical signal in human serum with a variance coefficient of 25.8% and a recovery of 93.6-110.6% for one-step subtype influenza virus detection.

Molybdenum Trioxide Quantum Dot-Encapsulated Nanogels for Virus Detection by Surface-Enhanced Raman Scattering on a 2D Substrate.

The use of nanogels (NGs) to modulate surface-enhanced Raman scattering (SERS) activities is introduced as an innovative strategy to address certain critical issues with SERS-based immunoassays. This includes the chemical deformation of SERS nanotags, as well as their nonspecific interactions and effective "hotspots" formation. Herein, the polymeric cocoon and stimuli-responsive properties of NGs were used to encapsulate SERS nanotags containing plasmonic molybdenum trioxide quantum dots (MoO3-QDs). The pH-controlled release of the encapsulated nanotags and their subsequent localization by maleimide-functionalized magnetic nanoparticles facilitated the creation of "hotspots" regions with catalyzed SERS activities. This approach resulted in developing a biosensing platform for the ultrasensitive immunoassays of hepatitis E virus (HEV) or norovirus (NoV). The immunoassays were optimized using the corresponding virus-like particles to attain limits of detection of 6.5 and 8.2 fg/mL for HEV-LPs and NoV-LPs, respectively. The SERS-based technique achieved a signal enhancement factor of up to ∼108 due to the combined electromagnetic and chemical mechanisms of the employed dual-SERS substrate of MoO3-QDs/2D hexagonal boron nitride nanosheets. The highlight and validation of the developed SERS-based immunoassays was the detection of NoV in infected patients' fecal specimen and clinical HEV G7 subtype. Importantly, this system can be used to maintain the stability of SERS nanotags and improve their reliability in immunoassays.

Plasmon Nanocomposite-Enhanced Optical and Electrochemical Signals for Sensitive Virus Detection.

The social impact of virus spread is immeasurable. Vaccine prophylaxes take considerable time to develop because clinical trials are required. The best initial response to an emerging virus is establishing a virus detection technology adapted by simply preparing virus-specific antibodies. A virus detection system that detects two signals from one analyte has been developed to detect the target virus more sensitively and reliably. Plasmon regions on the surface of nanoparticles are effective in enhancing optical and electrochemical signals. Thus, CdSeTeS quantum dots (QDs) have been used as optical and electrochemical signal-generating materials. In contrast, gold nanoparticle-magnetic nanoparticle-carbon nanotube (AuNP-MNP-CNT) nanocomposites are used for the magnetic separation of the virus from interferences and for signal enhancement. In the presence of the target virus, the QDs optically show a virus concentration-dependent fluorescence enhancement effect due to the localized surface plasmon resonance (LSPR) of AuNPs. Regarding the electrochemical signal, Cd ions eluted by acid degradation of the QDs in solution show a virus concentration-dependent increase in the current peak on an electrode whose electrochemical properties are improved by the deposition of these nanocomposites. Both nanomaterials are conjugated with antibodies specific to influenza virus A (IFV/A), binding this target in a sandwich structure. We are successfully detecting the virus from these two signals during actual virus detection, even when the virus particles are in a human serum matrix. The limit of detection is 2.16 fg/mL for optical detection and 13.66 fg/mL for electrochemical detection.

Self-assembled chromogen-loaded polymeric cocoon for respiratory virus detection.

Inspired by the self-assembly approach, in this work, the chromogen, 3,3',5,5'-tetramethylbenzidine (TMB), was successfully co-precipitated in aqueous solution to form collective nanoparticles (NPs) of signal molecules (TMB-NPs). Utilizing poly(lactide-co-glycolide) (PLGA) in the molecular delivery approach, the formed emulsion nanovesicle (TMB-NPs@PLGA) exhibits an enrichment of the collective signal molecules in a single antibody-antigen conjugation. A specific antibody-conjugated TMB-NPs@PLGA forms an immunocomplex sandwich structure upon the addition of influenza virus (IV)/A. The addition of dimethyl sulfoxide (DMSO) dissolves the PLGA nanovesicles, releasing the encapsulated TMB-NPs. Sequentially, the TMB-NPs release TMB molecules upon the addition of DMSO. The released TMB is catalytically oxidized by H2O2 with self-assembled protein-inorganic nanoflowers, where copper nanoflowers (CuNFs) acted as the nanozyme. The developed immunoassay demonstrates high sensitivity for IV/A with a limit of detection (LOD) as low as 32.37 fg mL-1 and 54.97 fg mL-1 in buffer and serum, respectively. For practical needs, a clinically isolated IV/A/H3N2 and spike protein of SARS-CoV-2 were detected with the LODs of 17 pfu mL-1 and 143 fg mL-1, respectively. These results show the applicability of the advanced TMB-NPs@PLGA-based colorimetric sensor for the highly sensitive detection of airborne respiratory viruses.

Hollow magnetic-fluorescent nanoparticles for dual-modality virus detection.

Combination of magnetic nanomaterials with multifunctionality is an emerging class of materials that exhibit tremendous potential in advanced applications. Synthesizing such novel nanocomposites without compromising magnetic behavior and introducing added functional properties is proven challenging. In this study, an optically active quantum dot (QD) (core) encapsulated inside iron oxide (hollow shell) is prepared as the first electrochemical/fluorescence dual-modality probe. Presence of magnetic layer on the surface enables excellent magnetic property and the encapsulating of QDs on the hollow shell structure maintains the fluorescence with minimal quenching effect, endowing for potential application with fluorescence modality readout. We successfully demonstrate dual-modality sensing utilizing of QD-encapsulated magnetic hollow sphere nanoparticles (QD@MHS NPs) with magnetic separation ability and highly integrated multimodal sensing for the detection of various viruses including hepatitis E virus (HEV), HEV-like particles (HEV-LPs), norovirus-like particles (NoV-LPs), and norovirus (NoV) from clinical specimens. Most importantly, fecal samples of HEV-infected monkey are successfully diagnosed with sensitivity similar to gold standard real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR). This well-defined QD@MHS NPs-based nanoplatform intelligently integrates dual-modality sensing and magnetic bio-separation, which open a gateway to provide an efficient point-of care testing for virus diagnostics.

Ultrasensitive Detection of the Hepatitis E Virus by Electrocatalytic Water Oxidation Using Pt-Co3O4 Hollow Cages.

A sensitive virus detection method applicable for an early stage increases the probability of survival. Here, we develop a simple and rapid detection strategy for the detection of the hepatitis E virus (HEV) by an electrocatalytic water oxidation reaction (WOR) using a platinum (Pt)-incorporated cobalt (Co)-based zeolite imidazole framework (ZIF-67). The surface cavity of ZIF-67 enables the rich loading of Pt NPs, and subsequent calcination etches the cavity, promoting the electrocatalytic activity of Pt-Co3O4 HCs. The Pt-Co3O4 HCs show excellent behavior for the WOR due to the synergistic interaction of Pt and Co3O4, evaluated by voltammetry and chronoamperometry. The synthesized Pt-Co3O4 HCs are conjugated with anti-HEV antibody (Ab@Pt-Co3O4 HCs); the electrocatalytic activity of Ab@Pt-Co3O4 HCs is combined with that of antibody-conjugated magnetic nanoparticles (MNPs) for HEV detection by a magneto-and-nanocomposite sandwich immunoassay. The sensor is challenged to detect the HEV in spiked serum samples and HEV G7 genotypes collected from the cell culture supernatant, reaching a low limit of detection down to 61 RNA copies mL-1. This work establishes a free-indicator one-step approach with the controlled design of Pt-Co3O4 HCs, which presents an effective WOR technique for virus detection in a neutral pH solution, which can be extended to electrocatalytic studies in the future integrated biosensing systems.

Plasmonic/magnetic molybdenum trioxide and graphitic carbon nitride quantum dots-based fluoroimmunosensing system for influenza virus.

A novel magnetic/plasmonic-assisted fluoro-immunoassay system is developed for the detection of influenza virus using magnetic-derivatized plasmonic molybdenum trioxide quantum dots (MP-MoO3 QDs) as the plasmonic/magnetic agent and fluorescent graphitic carbon nitride quantum dots (gCNQDs) as the monitoring probe. Specific antibody against influenza A virus was conjugated onto the surface of MP-MoO3 QDs and gCNQDs, respectively. In the presence of influenza A virus (as the test virus), a core-satellite immunocomplex is formed between the antibody-conjugated nanomaterials (Ab-MP-MoO3 QDs and Ab-gCNQDs) and their interaction resulted in the modulation and gradual enhancement of the fluorescence intensity of the detection probe with the influenza virus concentration-dependent increase. In addition, PL change without influenza A virus was not observed. Limits of detection of 0.25 and 0.9 pg/mL were achieved for Influenza virus A/New Caledonia (20/99/IVR/116) (H1N1) detection in deionized water and human serum, respectively. Clinically isolated influenza virus A/Yokohama (110/2009) (H3N2) was detected in the range of 45 - 25,000 PFU/mL, with a limit of detection ca 45 PFU/mL (as opposed to a minimum of 5000 PFU/mL for a commercial test kit). This developed biosensor provides a robust, sensitive as well as a selective platform for influenza virus detection.

Dual modality sensor using liposome-based signal amplification technique for ultrasensitive norovirus detection.

Sensitive and accurate detection methods for infectious viruses are the pressing need for effective disease diagnosis and treatment. Herein, based on V2O5 nanoparticles-encapsulated liposomes (VONP-LPs) we demonstrate a dual-modality sensing platform for ultrasensitive detection of the virus. The sensing performance relies on intrinsic peroxidase and electrochemical redox property of V2O5 nanoparticles (V2O5 NPs). The target-specific antibody-conjugated VONP-LPs and magnetic nanoparticles (MNPs) enrich the virus by magnetic separation and the separated VONP-LPs bound viruses are hydrolyzed to release the encapsulated V2O5 NPs. These released nanoparticles from captured liposomes act as peroxidase mimics and electrochemical redox indicator resulting in noticeable colorimetric and robust electrochemical dual-signal. Utilizing the superiority of dual-modality sensor with two quantitative analysis forms, norovirus like particles (NoV-LPs) can be detected by electrochemical signals with a wide linear range and low detection limit. To verify the applicability in real samples, norovirus (NoV) collected from actual clinical samples are effectively-identified with excellent accuracy. This proposed detection method can be a promising next-generation bioassay platform for early-stage diagnosis of virus disease and surveillance for public health.

Advancement of capture immunoassay for real-time monitoring of hepatitis E virus-infected monkey.

Rapid increasing outbreak of Hepatitis E virus (HEV) shows an urgent need of HEV detection. Instead of time consuming and expensive RT-qPCR, an efficient and quick monitoring system is in utmost demand which can be comparable with the RT-qPCR in term of reliability and detection limit. An advanced platform for immunoassay has been constructed in this study by a nanozyme that constitutes anti-HEV IgG antibody-conjugated gold nanoparticles (Ab-AuNPs) as core and in situ silver deposition on the surface of Ab-AuNPs as outer shell. The virus has been entrapped on the nanocomposites while the silver-shell has decomposed back to the silver ions (Ag+) by adding a tetramethylbenzidine (TMBZ) and hydrogen peroxide (H2O2) which indirectly quantifies the target virus concentration. Counterpart to only applying nanozyme, by incorporation of the enhanced effect of Ag shell on the AuNP-based nanozyme, the advance deposition has been confirmed to prove the signal amplification mechanism in the proposed immunoassay. Most importantly, the sensor performances have examined on the HEV, collected from the HEV-infected monkey over a period of 45 days. It was successfully correlated with the standard RT-qPCR data, showing the applicability of this immunoassay as a real-time monitoring on the HEV infection. The in situ formation of AuNPs@Ag as nanozyme in this capture immunoassay leads to a promising advancement over the conventional methods and nanozyme-based immunoassay in real application which can be a good substitute of RT-qPCR in near future.

Enhanced colorimetric detection of norovirus using in-situ growth of Ag shell on Au NPs.

Norovirus (NoV) is a leading cause of acute gastroenteritis. The low infectious dose and environmental stability of NoV facilitate its effective transmission through a variety of modes such as food, water and person-to-person. The available enzyme-linked immunosorbent assay (ELISA) for NoV detection has low sensitivity due to the low catalytic activity of the peroxidase used, and thus, a reliable ultrasensitive bioassay is needed. In this study, we apply the enhanced peroxidase-like activity of silver ion-incorporated gold nanoparticles (Au/Ag NPs) in a colorimetric bioassay for NoV detection. NoV was captured by anti-NoV genogroup II antibodies, which were immobilized on the surface of a 96-well microtiter plate and formed a sandwich structure among anti-NoV Ab, NoV and Ab-Au NP. Then, Ag ion-containing hydroquinone was added to form Au/Ag core/shell NPs. When H2O2/3,3',5,5'-tetramethylbenzidine (TMB) solution was added to the wells, Ag ions were liberated from the surface of Au/Ag NPs and enhanced the oxidation of TMB. These reactions enhanced the oxidation of TMB and developed an intense blue color. The Au/Ag NPs were shown to exhibit higher affinity and catalytic efficiency for H2O2 and higher catalytic velocity based on the kcat of up to 7-fold compared with Au NPs. The bioassay was then optimized to detect clinically isolated NoV using NoV-like particles (NoV-LPs). NoV-LPs were detected with a limit of detection of 10.8 pg/mL, corresponding to 1000- and 100-fold higher sensitivity compared to the gold-immunoassay and horseradish peroxidase-based ELISA, respectively. Clinically isolated NoV GII.4 and NoV GII.3 were detected in the range of 102-106 copies of viral RNA/mL fecal solution with a detection limit of 13.2 copies/mL fecal solution for NoV GII.4, equivalent to 132 copies of viral RNA/g feces and indicating significantly higher sensitivity compared to commercial immunoassay kits. This bioassay represents a workable detection assay for low concentrations of NoV that is applicable for early-stage diagnosis for public hygiene.

Femtomolar Detection of Dengue Virus DNA with Serotype Identification Ability.

Dengue surveillance trusts only on reverse transcription-polymerase chain reaction (RT-PCR) type methodologies for confirmation of dengue virus serotypes; however, its real time application is restricted due to the expensive, complicated, and time-consuming process. In search of a new sensing system, here, we have reported a two-way-detection method for Dengue virus (DENV) serotype identification along with DNA quantification by using a new class of nanocomposite of gold nanoparticles (AuNP) and nitrogen, sulfur codoped graphene quantum dots (N,S-GQDs). The N,S-GQDs@AuNP has been used for serotype detection via a simple fluorescence technique using four dye-combined probe DNAs which is further validated by confocal microscopy. The quantification of the DNA has been measured by the differential pulse voltammetric (DPV) technique using methyelene blue as a redox indicator. Results obtained in this study, clearly demonstrate that the N,S-GQDs@AuNP can efficiently detect the four serotypes of DENV individually in the concentration range of 10-14 to 10-6 M with the LOD of 9.4 fM. In addition, to confirm its applicability in long chained complex DNA system, the sensor was also applied to the clinically isolated DENV DNA and showed satisfactory performances for serotype identification as well as quantification. We hope this simple and reliable method can pave an avenue for the development of sensitive and robust sensing probes in biomedical applications.