NS1 glycoprotein detection in serum and urine as an electrochemical screening immunosensor for dengue and Zika virus.

The incidence of infection by the dengue virus (DENV) has grown dramatically, reaching 128 countries in tropical and subtropical regions worldwide, with a pattern of hyper-endemicity. DENV is a mosquito-borne disease having four serotypes, one or two circulating in epidemic outbreaks. The diagnosis of DENV is challenging mainly due to the circulation of new viruses with remarkable similarities, such as Zika (ZIKV) that may cause fetal microcephaly. DENV affects 390 million people per year, but these numbers may be higher due to the underreported and misclassified cases. Recently, the NS1 nonstructural protein has been described in serum and urine of DENV and ZIKV patients, suggesting its use as a biomarker for screening since a negative NS1 sample confirms the absence of these infections. Herein, a label-free immunosensor comprising an assembled nanostructured thin film of carbon nanotube-ethylenediamine is described. The advantage of in situ electrosynthesis of polymer film is to allow major control of thickness and conductivity, in addition to designing the reactive groups for functionalization. A quartz crystal microbalance system was used to estimate the thickness of the polymeric film obtained. The anti-NS1 monoclonal antibodies were immobilized to carbon nanotubes by covalent linkage, permitting a high stability during measurements. Analytical responses to NS1 were obtained by differential pulse voltammetry (DPV), showing a linear range from 20 to 800 ng mL-1 and reproducibility of 3.0%, with a limit of detection (LOD) of 6.8 ng mL- 1. This immunosensor was capable of detecting ZIKV and DENV NS1 in spiked urine and real serum in a clinical range.Graphical abstract.

Dengue immunoassay with an LSPR fiber optic sensor.

Dengue fever is a viral disease that affects millions of people worldwide. Specific tests for dengue are not usually performed due to high costs, complicated procedures and, in some cases, long time to yield a result. For widespread use of specific tests to be possible, fast, reliable and fairly simple methods are needed. In this paper, we present a new dengue diagnostic method for the acute phase of the infection. The method proposed uses an all-optical fiber sensor based on Localized Surface Plasmon Resonance (LSPR) and specular reflection from gold nanoparticles (AuNPs). Dengue anti-NS1 antibody was immobilized on AuNPs deposited on the endface of a standard multimode fiber (62.5 µm/125 µm). The sensor is able to detect NS1 antigen at different concentrations, with limit of quantification estimated to be 0.074 µg/ml = 1.54 nM. These results indicate that the sensor could potentially be used for dengue diagnosis in the acute phase of the infection.

A simple HPV 18 detection method based on ultra specific primer immobilized on glass slides.

This study was carried out to develop a simple and inexpensive method for detection of Human papillomavirus (HPV 18) based on irreversible immobilization of ultra specific primer on silanized glass slides. This method is revealed by Blue Green Loading Dye I (LGC) and compared with conventional polymerase chain reaction (PCR) for endocervical samples. The new method was tested in 40 DNA samples with precancer uterine lesions of women treated in Hospital of Recife PE, Brazil. DNA samples were extracted using Wizard Genomic DNA Purification Kit according to manufacturer's instructions. The samples were tested for HPV 18 by conventional PCR (PCRc) and the products visualized on 1.2% agarose gel, with LGC under UV 260 nm. After that the positive and negative samples to HPV 18 were tested by immobilization method and the results visualized with LGC under UV 260 nm. Both PCRc and immobilization method showed high degree of correlation (95%), whereas comparison between PCRc and immobilization method showed good correlation (100%). PCRc is widely known for detection of HPV because of its high sensitivity and efficiency, but due to high cost it is not yet standardized for use in public health laboratories. In our study, the single-stranded DNA immobilized method on a glass slide was effective in screening for HPV revealed by Blue Green and may be an alternative method for diagnosis of HPV once it offers a fast, and easy handling.

An ultrasensitive electrochemical immunosensor for hepatitis C antibodies based on one-step-eletrosynthetized polypyrrole-graphene nanocomposite.

An ultrasensitive label-free electrochemical immunosensor was developed for hepatitis C antibodies (anti-HCV). Worldwide, it is estimated 71 million people have HCV infection in a chronic stage that may lead to cirrose and cancer. To achieve HCV elimination, health programs should include screening testing based on anti-HCV detection allowing the early-stage treatment. The immunosensor was based on a graphene oxide-polypyrrole (PPy-GO) film one-step electropolymerized on the electrode surface. Ultrasensitive anti-HCV detection was ensured by HCV antigen conjugated to biotin that was immobilized in a great amount on streptavidin-coated nanostructured surface. Analytical responses were obtained by anodic peaks from the square wave voltammetry in the presence of ferrocyanide/ferricyanide as a redox probe. This immunosensor exhibited a linear range from 2 to 14 ng mL-1 of anti-HCV and a limit of detection in the clinical range (1.63 ng mL-1). Furthermore, the immunosensor presented an efficient performance for the determination of anti-HCV in spiked serum samples, becoming this developed nanosensor as potential tools for early HCV diagnosis and screening. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10853-022-06992-5.

A Polypyrrole/Nanoclay Hybrid Film for Ultra-Sensitive Cardiac Troponin T Electrochemical Immunosensor.

An electrochemical immunosensor based on a nanohybrid film of carboxylated polypyrrole and amine nanoclay was developed for label-free detection of the human cardiac troponin T (cTnT). The nanohybrid film was formed in situ on the surface of the glassy carbon electrode, followed by the covalent immobilization of anti-troponin T antibodies by glutaraldehyde. Morphological and chemical characterizations of the nanohybrid film were performed by scanning electron microscopy and Fourier-transform infrared spectroscopy. Under the optimized conditions, a calibration curve for cTnT in spiked serum was obtained by square wave voltammetry, and a low limit of detection and quantification was achieved (0.35 and 1.05 pg mL-1, respectively). This was the first time that this type of nanohybrid film was used in the development of an immunosensor for cTnT that proved to be a simple and efficient strategy for the manufacture of a label-free electrochemical device that could be applied in the diagnosis of acute myocardial infarction.

A piezoelectric immunosensor for Leishmania chagasi antibodies in canine serum.

The American visceral leishmaniasis is an important cause of morbidity and mortality in Brazil for both humans and dogs. Attempts to make a diagnosis of this disease need to be improved, especially in endemic areas, and in the tracking and screening of asymptomatic dogs, which are their main host in urban areas. A quartz crystal microbalance immunosensor for the diagnosis of the canine visceral leishmaniasis using a recombinant antigen of Leishmania chagasi (rLci2B-NH6) was developed. The rLci2B-NH6 was tightly immobilized on a quartz crystal gold electrode by self-assembled monolayer based on short-chain length thiol. The strategy was the use of the antigen-histidine tail covalently linked to glutaraldehyde performing a Schift base which permits a major exposure of epitopes and a reduced steric hindrance. The immunosensor showed good results regarding sensitivity and reproducibility, being able to distinguish positive and negative canine serum for L. chagasi. Furthermore, the immunosensor can be reused through exposure to sodium dodecyl sulfate solution, which promotes the dissociation of antigen-antibody binding, restoring the sensor surface with immobilized biologically active antigens for further analysis.

A nanostructured piezoelectric immunosensor for detection of human cardiac troponin T.

A piezoelectric immunosensor based on gold nanoparticles (AuNPs) co-immobilized on a dithiol-modified surface is proposed for detection of human cardiac troponin T (TnT). Anti-human troponin T (anti-TnT) antibodies were covalently immobilized on the nanostructured electrode surface by thiol-aldehyde linkages. In a homogeneous bulk solution, TnT was captured by anti-TnT immobilized on the QCM electrode. Cyclic voltammetry studies were used to characterize the AuNPs layer on the electrode surface and the anti-TnT immobilization steps. The QCM-flow immunosensor exhibited good reliability, measuring concentrations of TnT from 0.003 to 0.5 ng mL(-1) in human serum with high linearity (r = 0.989; p < 0.01). The immunosensor exhibited a 7% coefficient of variation and 0.0015 ng mL(-1) limit of detection, indicating a high reproducibility and sensitivity. The proposed QCM nanostructured immunosensor is easy to use and has promising potential in the diagnosis of acute myocardial infarction due to its speed and high sensitivity.

Plastic Antibody of Polypyrrole/Multiwall Carbon Nanotubes on Screen-Printed Electrodes for Cystatin C Detection.

This work reports the design of a novel plastic antibody for cystatin C (Cys-C), an acute kidney injury biomarker, and its application in point-of-care (PoC) testing. The synthetic antibody was obtained by tailoring a molecularly imprinted polymer (MIP) on a carbon screen-printed electrode (SPE). The MIP was obtained by electropolymerizing pyrrole (Py) with carboxylated Py (Py-COOH) in the presence of Cys-C and multiwall carbon nanotubes (MWCNTs). Cys-C was removed from the molecularly imprinted poly(Py) matrix (MPPy) by urea treatment. As a control, a non-imprinted poly(Py) matrix (NPPy) was obtained by the same procedure, but without Cys-C. The assembly of the MIP material was evaluated in situ by Raman spectroscopy and the binding ability of Cys-C was evaluated by the cyclic voltammetry (CV) and differential pulse voltammetry (DPV) electrochemical techniques. The MIP sensor responses were measured by the DPV anodic peaks obtained in the presence of ferro/ferricyanide. The peak currents decreased linearly from 0.5 to 20.0 ng/mL of Cys-C at each 20 min successive incubation and a limit of detection below 0.5 ng/mL was obtained at pH 6.0. The MPPy/SPE was used to analyze Cys-C in spiked serum samples, showing recoveries <3%. This device showed promising features in terms of simplicity, cost and sensitivity for acute kidney injury diagnosis at the point of care.

A Label and Probe-Free Zika Virus Immunosensor Prussian Blue@carbon Nanotube-Based for Amperometric Detection of the NS2B Protein.

Zika virus (ZIKV) is a mosquito-borne infection, predominant in tropical and subtropical regions causing international concern due to the ZIKV disease having been associated with congenital disabilities, especially microcephaly and other congenital abnormalities in the fetus and newborns. Development of strategies that minimize the devastating impact by monitoring and preventing ZIKV transmission through sexual intercourse, especially in pregnant women, since no vaccine is yet available for the prevention or treatment, is critically important. ZIKV infection is generally asymptomatic and cross-reactivity with dengue virus (DENV) is a global concern. An innovative screen-printed electrode (SPE) was developed for amperometric detection of the non-structural protein (NS2B) of ZIKV by exploring the intrinsic redox catalytic activity of Prussian blue (PB), incorporated into a carbon nanotube-polypyrrole composite. Thus, this immunosensor has the advantage of electrochemical detection without adding any redox-probe solution (probe-less detection), allowing a point-of-care diagnosis. It was responsive to serum samples of only ZIKV positive patients and non-responsive to negative ZIKV patients, even if the sample was DENV positive, indicating a possible differential diagnosis between them by NS2B. All samples used here were confirmed by CDC protocols, and immunosensor responses were also checked in the supernatant of C6/36 and in Vero cell cultures infected with ZIKV.

Non-structural protein 1 from Zika virus: Heterologous expression, purification, and potential for diagnosis of Zika infections.

Zika virus (ZIKV) infections were associated with neurological disorders only after the Brazilian outbreak in 2015. The lack of vaccines and precise diagnosis requires a precise method to detect ZIKV infection. This study aimed to evaluate three ZIKV recombinant proteins for the development of ZIKV infections. Here, it was purified stable recombinant ZIKV Capsid (r-ZIKV-c), non-structural proteins NS1 (r-ZIKV-NS1), and NS3 (r-ZIKV-NS3) for detection of the infection by ZIKV in blood sera of patients. A commercial polyclonal antibody recognized the r-ZIKV-NS1. Here, among three proteins, NS1 showed the best result for diagnostic purposes using serum samples, despite the high similarity with NS1 from DENV, and could differentiate the infections. The recombinant NS1 was used to produce a monoclonal antibody to differentiate between DENV and ZIKV NS1. As for recombinant proteins, the result for r-ZIKV-NS1 values showed 77% and 100% sensitivity and specificity, respectively, in the IgM assay. Our data showed the protein could successfully differentiate between sera of ZIKV infected patients from sera of those not infected with the virus and differentiate from sera of DENV infected patients. Thus, the generated recombinant proteins have great potential for serological diagnosis of ZIKV in Brazil, where it is indispensable.

An ultrasensitive Cystatin C renal failure immunosensor based on a PPy/CNT electrochemical capacitor grafted on interdigitated electrode.

An interdigitated immunosensor for Cystatin C detection based on polypyrrole/carbon nanotube electrochemical capacitor is described. Cystatin C (CysC) is powerful biomarker for early acute renal failure and one predictive for cardiovascular risk, sepsis, cancer and death. Recently, electrochemical immunosensors based on interdigitated electrodes (IDE) have been successfully focused on development of point-of-care testing, due to their miniaturization facilities and higher sensitivity as compared with the screen-printed electrochemical sensing. Herein, a polypyrrole/carbon nanotube nanoyhibrid film was grafted on two gold fingers by electropolymerization obtaining a supercapacitor. Anti-CysC antibodies were immobilized on the IDE by covalent entrapment via ethylenediamine bifunctional agent, followed by glycine blocking in acid and alkaline medium. Under low frequency, capacitive effect of antigen-antibody interaction were observed by double layer capacitance, and analytical responses of this IDE immunosensor to CysC serum were obtained by changes on phase angle a linear range up to 300 ng/mL. The cutoff was calculated for serum samples showing a total reducing of non-specific binding at approximately 28 ng/mL CysC. This immunosensor based on interdigitated electrode (IDE) is a potential tools as portable device,with possibility to use as a practical and rapid test for CysC diagnostic in samples of serum.

A probeless and label-free electrochemical immunosensor for cystatin C detection based on ferrocene functionalized-graphene platform.

A novel electrochemical sensor with inherent redox activity mediated by ferrocene for Cystatin C (CysC), an early kidney failure biomarker, is described. The current response was mediated by graphene oxide-ferrocene nanofilm with redox-activity coming from electroactive species surface-confined. Anti-CysC antibodies were immobilized by their Fc portions on the drop-casting polyethyleneimine (PEI) film for improving the sensitivity and reproducibility. Stepwise modifications of the nanostructured surface were characterized by electrochemical techniques, FT-IR and AFM. FT-IR confirmed the formation of the Fc-GO nanocomposite and PEI deposition on the electrode surface. The AFM micrographs confirmed a nanometric film of Fc-GO and PEI. The sensor platform showed a response from 0.1 to 1000 ng/mL and lower limit of detection (LOD) of 0.03 ng/mL of CysC, with good accuracy, specificity and it was successfully applied for CysC detection. Advantages of this immunosensor include rapid testing with minimal steps by the simple use of an intrinsic redox probe, working in a reduction potential, which avoids potential interferences. This proposal attempts to circumvent amperometric detection limitations and provides a promising candidate for future point-of-care diagnostics without redox probe additional solutions for measurements.

A label-free and reagentless immunoelectrode for antibodies against hepatitis B core antigen (anti-HBc) detection.

An electrochemical immunosensor devoted the core hepatitis B antibody detection, based on polytyramine (PTy) and carbon nanotubes (CNT) composite was developed. The antibody interactions with immobilized antigens were detected by reduction on the electron transfer from ionic species coming from reactive amine groups of PTy. The synthesis in acid medium of PTy-CNT composite favorite a great amount of NH3+ ionic species, forming a nanocomposite with high catalytic activity on the electrode surface. As proof-of-concept, antibodies against the core hepatitis B virus were label-free and reagentless electrochemically detected by square wave voltammetry (SWV) through decrease on cathodic peaks. It was recently reported that hepatitis B core antigen antibodies (anti-HBc) is a powerful biomarker for hepatitis B virus (HBV) infection, being more specific than HBsAg due to the possibility of detecting the occult HBV infections. The nanostructured film was characterized by atomic force microscopy and electrochemical techniques. This immunosensor showed linear responses from 1.0 to 5.0 ng mL-1 and a limit of detection of 0.89 ng mL-1 anti-HBc. It was also tested in assays with negative and positive blood samples using 0.1 M KCl as electrolyte support on readings showing specific responses. This easy reagentless detection platform, showing a remarkable potential to development of bolder and simpler HBV assay for screening of blood bags, in attempting to circumvent point-of-care testing limitations.

Electrochemical immunosensor for differential diagnostic of Wuchereria bancrofti using a synthetic peptide.

Lymphatic filariasis (LF) is a neglected tropical disease transmitted by mosquitoes and the second cause of permanent disability leading to a significant morbidity and mortality rate. Previously, we have identified epitopes of the filarial abundant larval transcript-2 (ALT-2) protein using a microarray mapping. In this study, one of the epitopes (Wb/ALT2-A5) was used to construct an electrochemical immunosensor. Electrochemical technique of cyclic voltammetry was performed for detecting the signal generated by the interaction between the (Wb/ALT2-A5) peptide and circulating antibodies of serum human samples. (Wb/ALT2-A5) epitope antigens were successfully immobilized on the working electrode of a screen-printed carbon electrode (SPCE) by their amine groups via chitosan film by coupling with glutaraldehyde as crosslinker. After the sensor ready, a pool of human sera infected with Wuchereria bancrofti was added to its surface. Electrochemical responses were generated by applying a potential of - 0.6 to 0.6 V, scan rate of 0.025 V/s. A detection limit of 5.0 µg mL-1 for the synthetic peptides (Wb/ALT2-A5) and 0.002 µg mL-1 for human serum, with a sensitivity of 1.86 µA. The performance of this assay was successfully tested in human serum samples from infected and healthy patients. Thus, this proposed immunosensor, which is able to identify circulating antibodies, can be applied to the diagnosis of the W. bancrofti parasitic disease.

A label-free electrochemical immunosensor for hepatitis B based on hyaluronic acid-carbon nanotube hybrid film.

An electrochemical immunosensor developed for detection of antibodies to hepatitis B core protein (anti-HBc) is described. Anti-HBc is the earliest serological marker from hepatitis B virus (HBV) infection, remaining all life after contact with virus, being considered the most important marker for uses in screening of blood bank. A nanohybrid surface assembled onto a glassy carbon electrode consisting of amino carbon nanotubes recovered by hyaluronic acid was used as sensing platform to detect the anti-HBc. All the steps of electrode surface modification were characterized by Scanning Electronic Microscopy and extensively evaluated by electrochemical techniques. The electrode response was measured by direct anti-HBc antigen interactions by square wave voltammetry, dispensing uses of label or chemical mediators. Under optimal conditions, the anodic peak current which was proportional to the anti-HBs concentration. The immunosensor response was linear toward anti-HBc in concentrations up to 6 ng mL(-1), with a detection limit of 0.03 ng mL(-1). The linear range achieved was according to clinical level, indicating the immunosensor as promising tool for use as a criterion for blood bag disposal. The enhancement of the hyaluronic acid by carbon nanotube promoted an increase of charge electron transfer, besides a stable platform for HBc.

An ultrasensitive human cardiac troponin T graphene screen-printed electrode based on electropolymerized-molecularly imprinted conducting polymer.

A nano-molecularly imprinted polymer (N-MIP) assembled on a screen-printed electrode for the cardiac troponin T (cTnT) was developed. The biomimetic surface was obtained by a co-polymer matrix assembled on the reduced graphene oxide (RGO) electrode surface. The cTnT active sites were engineered using pyrrole and carboxylated pyrrole that was one-step electropolymerized jointly with cTnT by cyclic voltammetry. The stepwise preparation of the biomimetic surface was characterized by cyclic and differential pulse voltammetries using the ferrocyanide/ferricyanide as redox probe. Structural and morphological characterization was also performed. The optimal relation of pyrrole and pyrrole-3-acid carboxylic to perform the cTnT biomimetic nanosurface was obtained at 1:5 ratio. The analytical performance of cTnT N-MIP performed by differential pulse voltammetry showed a linear range from 0.01 to 0.1 ngmL(-1) (r=0.995, p«0.01), with a very low limit of detection (0.006 ngmL(-1)). The synergic effect of conductive polymer and graphene forming 3D structures of reactive sites resulted in a N-MIP with excellent affinity to cTnT binding (KD=7.3 10(-13) molL(-1)). The N-MIP proposed is based on a simple method of antibody obtaining with a large potential for point-of-care testing applications.

Cratylia mollis lectin nanoelectrode for differential diagnostic of prostate cancer and benign prostatic hyperplasia based on label-free detection.

The research for new biomarkers of cancer has studied the role of fetuin glycoprotein on the metastatic disease diagnosis. Cratylia mollis is a lectin with high finity to fetuin, and used here to differentiate prostate cancer and benign prostatic hyperplasia. A label-free electrochemical nanosensor based on assembled carboxylated carbon nanotubes (COOH-CNTs) and poly-L-lysine (PLL) film was developed and applied to serum samples of prostate cancer positive for Gleason score. The electrode analytical response to fetuin in PBS samples, obtained by square wave voltammetry, exhibited a linear range from 0.5 to 25µgmL(-1), with a high correlation coefficient (r=0.994, p<0.001) and low limit of detection (0.017µgmL(-1)). The lectin nanoelectrode showed a good repeatability (1.24% RSD) and reproducibility (4.24% RSD). A pool of serum samples from prostate cancer patients with known the Gleason score were tested showing a significant statistically correlation. Thus, the lectin nanoelectrode was able to distinguish the degree of staging prostate cancer, providing the diagnostic differentiation of benign and malign hyperplasia. To the best of our knowledge, it is the first biosensor for this application using a lectin.

Amino-functionalization of carbon nanotubes by using a factorial design: human cardiac troponin T immunosensing application.

A simple amino-functionalization method for carbon nanotubes and its application in an electrochemical immunosensor for detection of the human cardiac troponin T are described. Amino-functionalized carbon nanotubes allow oriented antibodies immobilization via their Fc regions, improving the performance of an immunosensor. Herein multiwalled carbon nanotubes were amino-functionalized by using the ethylenediamine reagent and assays were designed by fractional factorial study associated with Doehlert matrix. Structural modifications in the carbon nanotubes were confirmed by Fourier transform infrared spectroscopy. After amino-functionalization the carbon nanotubes were attached to screen-printed carbon electrode and a sandwich-type immunoassay was performed for measuring the cardiac troponin T. The electrochemical measurements were obtained through hydrogen peroxide reaction with peroxidase conjugated to the secondary antibody. Under optimal conditions, troponin T immunosensor was evaluated in serum samples, which showed a broad linear range (0.02 to 0.32 ng mL(-1)) and a low limit of detection, 0.016 ng mL(-1). This amino platform can be properly used as clinical tool for cardiac troponin T detection in the acute myocardial infarction diagnosis.

A carbon nanotube screen-printed electrode for label-free detection of the human cardiac troponin T.

Label-free immunosensor based on amine-functionalized carbon nanotubes screen-printed electrode is described for detection of the cardiac troponin T, an important marker of acute myocardial infarction. The disposable sensor was fabricated by tightly squeezing an adhesive carbon ink containing carbon nanotubes onto a polyethylene terephthalate substrate forming a thin film. The use of carbon nanotubes increased the reproducibility and stability of the sensor, and the amine groups permitted nonrandom immobilization of antibodies against cardiac troponin T. Amperometric responses were obtained by differential pulse voltammetry in presence of a ferrocyanide/ferricyanide redox probe after troponin T incubation. The calibration curve indicated a linear response of troponin T between 0.0025 ng mL(-1) and 0.5 ng mL(-1), with a good correlation coefficient (r=0.995; p<0.0001, n=7). The limit of detection (0.0035 ng mL(-1) cardiac troponin T) was lower than any previously described by immunosensors and was comparable with conventional analytical methods. The high reproducibility and clinical range obtained using this immunosensor support its utility as a potential tool for point-of-care acute myocardial infarction diagnostic testing.

A sensor tip based on carbon nanotube-ink printed electrode for the dengue virus NS1 protein.

An immunosensor for the non-structural protein 1 (NS1) of the dengue virus based on carbon nanotube-screen printed electrodes (CNT-SPE) was successfully developed. A homogeneous mixture containing carboxylated carbon nanotubes was dispersed in carbon ink to prepare a screen printed working electrode. Anti-NS1 antibodies were covalently linked to CNT-SPE by an ethylenediamine film strategy. Amperometrical responses were generated at -0.5 V vs. Ag/AgCl by hydrogen peroxide reaction with peroxidase (HRP) conjugated to the anti-NS1. An excellent detection limit (in the order of 12 ng mL(-1)) and a sensitivity of 85.59 µA mM(-1)cm(-2) were achieved permitting dengue diagnostic according to the clinical range required. The matrix effect, as well as the performance of the assays, was successfully evaluated using spiked blood serum sample obtaining excellent recovery values in the results. Carbon nanotubes incorporated to the carbon ink improved the reproducibility and sensitivity of the CNT-SPE immunosensor. This point-of-care approach represents a great potential value for use in epidemic situations and can facilitate the early screening of patients in acute phase of dengue virus.