RESUMO

Self-healing materials inspire the next generation of multifunctional wearables and Internet of Things appliances. They expand the realm of thin film fabrication, enabling seamless conformational coverage irrespective of the shape complexity and surface geometry for electronic skins, smart textiles, soft robotics, and energy storage devices. Within this context, the layer-by-layer (LbL) technique is versatile for homogeneously dispersing materials onto various matrices. Moreover, it provides molecular level thickness control and coverage on practically any surface, with poly(ethylenimine) (PEI) and poly(acrylic acid) (PAA) being the most used materials primarily employed in self-healing LbL structures operating at room temperature. However, achieving thin film composites displaying controlled conductivity and healing ability is still challenging under ambient conditions. Here, PEI and PAA are mixed with conductive fillers (gold nanorods, poly(3,4-ethylene dioxythiophene): polystyrenesulfonate (PEDOT:PSS), reduced graphene oxides, and multiwalled carbon nanotubes) in distinct LbL film architectures. Electrical (AC and DC), optical (Raman spectroscopy), and mechanical (nanoindentation) measurements are used for characterizing composite structures and properties. A delicate balance among electrical, mechanical, and structural characteristics must be accomplished for a controlled design of conductive self-healing composites. As a proof-of-concept, four LbL composites were chosen as sensing units in the first reported self-healing e-tongue. The sensor can easily distinguish basic tastes at low molar concentrations and differentiate trace levels of glucose in artificial sweat. The formed nanostructures enable smart coverages that have unique features for solving current technological challenges.

RESUMO

SARS-CoV-2 rapid spread required urgent, accurate, and prompt diagnosis to control the virus dissemination and pandemic management. Several sensors were developed using different biorecognition elements to obtain high specificity and sensitivity. However, the task to achieve these parameters in combination with fast detection, simplicity, and portability to identify the biorecognition element even in low concentration remains a challenge. Therefore, we developed an electrochemical biosensor based on polypyrrole nanotubes coupled via Ni(OH)2 ligation to an engineered antigen-binding fragment of heavy chain-only antibodies (VHH) termed Sb#15. Herein we report Sb#15-His6 expression, purification, and characterization of its interaction with the receptor-binding domain (RBD) of SARS-CoV-2 in addition to the construction and validation of a biosensor. The recombinant Sb#15 is correctly folded and interacts with the RBD with a dissociation constant (KD) of 27.1 ± 6.4 nmol/L. The biosensing platform was developed using polypyrrole nanotubes and Ni(OH)2, which can properly orientate the immobilization of Sb#15-His6 at the electrode surface through His-tag interaction for the sensitive SARS-CoV-2 antigen detection. The quantification limit was determined as 0.01 pg/mL using recombinant RBD, which was expressively lower than commercial monoclonal antibodies. In pre-characterized saliva, both Omicron and Delta SARS-CoV-2 were accurately detected only in positive samples, meeting all the requirements recommended by the World Health Organization for in vitro diagnostics. A low sample volume of saliva is needed to perform the detection, providing results within 15 min without further sample preparations. In summary, a new perspective allying recombinant VHHs with biosensor development and real sample detection was explored, addressing the need for accurate, rapid, and sensitive biosensors.

RESUMO

An innovative strategy is proposed to simultaneously exfoliate multi-walled carbon nanotubes (MWCNTs) and generate MWCNTs with immunoaffinity properties. This strategy was based on the non-covalent functionalization of MWCNTs with human immunoglobulin G (IgG) by sonicating 2.5 mg mL-1 MWCNTs in 2.0 mg mL-1 IgG for 15 min with sonicator bath. Impedimetric experiments performed at glassy carbon electrodes (GCE) modified with the resulting MWCNT-IgG nanohybrid in the presence of anti-human immunoglobulin G antibody (Anti-IgG) demonstrated that the immunoglobulin retains their biorecognition properties even after the treatment during the MWCNT functionalization. We proposed, as proof-of-concept, two model electrochemical sensors, a voltammetric one for uric acid quantification by taking advantages of the exfoliated MWCNTs electroactivity (linear range, 5.0 × 10-7 M - 5.0 × 10-6 M; detection limit, 165 nM) and an impedimetric immunosensor for the detection of Anti-IgG through the use of the bioaffinity properties of the IgG present in the nanohybrid (linear range, 5-50 µg mL-1; detection limit, 2 µg mL-1).

Assuntos

RESUMO

In this study, polypyrrole nanotubes (PPy-NT) and gold nanoparticles (AuNPs) were electrochemically synthesized to form a hybrid material and used as an electroactive layer for the attachment of proteins for the construction of a high-performance biosensor. Besides the enhancement of intrinsic conductivity of the PPy-NT, the AuNPs act as an anchor group for the formation of self-assembly monolayers (SAMs) from the gold-sulfur covalent interaction between gold and Mercaptopropionic acid (MPA). This material was used to evaluate the viability and performance of the platform developed for biosensing, and three different biological approaches were tested: first, the Avidin-HRP/Biotin couple and characterizations were made by using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), wherein we detected Biotin in a linear range of 100-900 fmol L-1. The studies continued with folate group biomolecules, using the folate receptor α (FR-α) as a bioreceptor. Tests with anti-FR antibody detection were performed, and the results obtained indicate a linear range of detection from 0.001 to 6.70 pmol L-1. The same FR-α receptor was used for Folic Acid detection, and the results showed a limit of detection of 0.030 nmol L-1 and a limit of quantification of 90 pmol L-1. The results indicate that the proposed biosensor is sensitive and capable of operating in a range of clinical interests.

Assuntos

RESUMO

We report the advantages of glassy carbon electrodes (GCE) modified with multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with polyarginine (PolyArg) for the adsorption and electrooxidation of different DNAs and the analytical applications of the resulting platform. The presence of the carbon nanostructures, and mainly the charge of the PolyArg that supports them, facilitates the adsorption of calf-thymus and salmon sperm double-stranded DNAs and produces an important decrease in the overvoltages for the oxidation of guanine and adenine residues and a significant enhancement in the associated currents. As a proof-of-concept of possible GCE/MWCNTs-PolyArg biosensing applications, we develop an impedimetric genosensor for the quantification of microRNA-21 at femtomolar levels, using GCE/MWCNTs-PolyArg as a platform for immobilizing the DNA probe, with a detection limit of 3fM, a sensitivity of 1.544 × 103 Ω M-1, and a successful application in enriched biological fluids.

RESUMO

We report two novel genosensors for the quantification of SARS-CoV-2 nucleic acid using glassy carbon electrodes modified with a biocapture nanoplatform made of multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with avidin (Av) as a support of the biotinylated-DNA probes. One of the genosensors was based on impedimetric transduction offering a non-labelled and non-amplified detection of SARS-CoV-2 nucleic acid through the increment of [Fe(CN)6]3-/4- charge transfer resistance. This biosensor presented an excellent analytical performance, with a linear range of 1.0 × 10-18 M - 1.0 × 10-11 M, a sensitivity of (5.8 ± 0.6) x 102 Ω M-1 (r2 = 0.994), detection and quantification limits of 0.33 aM and 1.0 aM, respectively; and reproducibilities of 5.4% for 1.0 × 10-15 M target using the same MWCNTs-Av-bDNAp nanoplatform, and 6.9% for 1.0 × 10-15 M target using 3 different nanoplatforms. The other genosensor was based on a sandwich hybridization scheme and amperometric transduction using the streptavidin(Strep)-biotinylated horseradish peroxidase (bHRP)/hydrogen peroxide/hydroquinone (HQ) system. This genosensor allowed an extremely sensitive quantification of the SARS-CoV-2 nucleic acid, with a linear range of 1.0 × 10-20 M - 1.0 × 10-17 M, detection limit at zM level, and a reproducibility of 11% for genosensors prepared with the same MWCNTs-Av-bDNAp1 nanoplatform. As a proof-of-concept, and considering the extremely high sensitivity, the genosensor was challenged with highly diluted samples obtained from SARS-CoV-2 RNA PCR amplification.

RESUMO

Coronavirus disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is considered one of the worst pandemic outbreaks worldwide. This ongoing pandemic urgently requires rapid, accurate, and specific testing devices to detect the virus. We report a simple electrochemical biosensor based on a highly specific synthetic peptide to detect SARS-CoV-2 Spike protein. Unlike other reported electrochemical biosensors involving nanomaterials or complex approaches, our electrochemical platform uses screen-printed gold electrodes functionalized with the thiolated peptide, whose interaction with the Spike protein is directly followed by Electrochemical Impedance Spectroscopy. The electrochemical platform was Spike protein concentration-dependent, with high sensitivity and reproducibility and a limit of detection of 18.2 ng/mL when tested in Spike protein commercial solutions and 0.01 copies/mL in lysed SARS-CoV-2 particles. The label-free biosensor successfully detected the Spike protein in samples from infected patients straightforwardly in only 15 min. The simplicity of the proposed format combined with an on-demand designed peptide opens the path for detecting other pathogen-related antigens.

Assuntos

RESUMO

The rapid and reliable detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroconversion in humans is crucial for suitable infection control. In this sense, many studies have focused on increasing the sensibility, lowering the detection limits and minimizing false negative/positive results. Thus, biosensors based on nanoarchitectures of conducting polymers are promising alternatives to more traditional materials since they can hold improved surface area, higher electrical conductivity and electrochemical activity. In this work, we reported the analytical comparison of two different conducting polymers morphologies for the development of an impedimetric biosensor to monitor SARS-CoV-2 seroconversion in humans. Biosensors based on polypyrrole (PPy), synthesized in both globular and nanotubular (NT) morphology, and gold nanoparticles are reported, using a self-assembly monolayer of 3-mercaptopropionic acid and covalently linked SARS-CoV-2 Nucleocapsid protein. First, the novel hybrid materials were characterized by electron microscopy and electrochemical measurements, and the biosensor step-by-step construction was characterized by electrochemical and spectroscopic techniques. As a proof of concept, the biosensor was used for the impedimetric detection of anti-SARS-CoV-2 Nucleocapsid protein monoclonal antibodies. The results showed a linear response for different antibody concentrations, good sensibility and possibility to quantify 7.442 and 0.4 ng/mL of monoclonal antibody for PPy in the globular and NT morphology, respectively. The PPy-NTs biosensor was able to discriminate serum obtained from COVID-19 positive versus negative clinical samples and is a promising tool for COVID-19 immunodiagnostic, which can contribute to further studies concerning rapid, efficient, and reliable detections.

RESUMO

Electrochemical sensors and biosensors are useful techniques for fast, inexpensive, sensitive, and easy detection of innumerous specimen. In face of COVID-19 pandemic, it became evident the necessity of a rapid and accurate diagnostic test, so the impedimetric immunosensor approach can be a good alternative to replace the conventional tests due to the specific antibody-antigen binding interaction and the fast response in comparison to traditional methods. In this work, a modified electrode with electrosynthesized PEDOT and gold nanoparticles followed by the immobilization of truncated nucleoprotein (N aa160-406aa) was used for a fast and reliable detection of antibodies against COVID-19 in human serum sample. The method consists in analyzing the charge-transfer resistance (RCT) variation before and after the modified electrode comes into contact with the positive and negative serum sample for COVID-19, using [Fe(CN)6]3-/4- as a probe. The results show a linear and selective response for serum samples diluted in a range of 2.5 × 103 to 20 × 103. Also, the electrode material was fully characterized by Raman spectroscopy, transmission electron microscopy and scanning electron microscopy coupled with EDS, indicating that the gold nanoparticles were well distributed around the polymer matrix and the presence of the biological sample was confirmed by EDS analysis. EIS measurements allowed to differentiate the negative and positive samples by the difference in the RCT magnitude, proving that the material developed here has potential properties to be applied in impedimetric immunosensors for the detection of SARS-CoV-2 antibodies in about 30 min.

RESUMO

Dengue is one of the most commonly neglected tropical diseases transmitted by Aedes aegypti infected with Dengue virus. This virus belongs to the gender Flavivirus and produces a non-structural protein 1 (NS1), which is an important biomarker found at high levels in blood in early disease stage. Therefore, this study focused on the development of an electrochemical biosensor for NS1 detection using DNA aptamers. Gold electrodes were co-immobilized with specific aptamers and 6-mercapto-1-hexanol (MCH) to obtain a self-assembled monolayer. The molar ratio between aptamers and MCH was optimized and the platform characterized by electrochemical impedance spectroscopy and atomic force microscopy. Bovine serum albumin was added in NS1 solution to stabilize it and block the surface to avoid non-specific interactions. The biosensor performance was tested with NS1 protein serotype 4 (in phosphate saline buffer and human serum) and with a solution of serotype 1 in human serum. The results showed a sensitivity of 2.9%, 2.7% and 1.7% per decade, respectively, and low limit of detection (0.05, 0.022 and 0.025 ng/mL). The platform was also tested with Envelope protein as negative control. Furthermore, the aptamer sensor was able to detect NS1 in clinical range and it is a promising candidate for a new class for miniaturized point-of-care device for different Dengue serotypes.

Assuntos

RESUMO

Breast cancer remains one of the leading causes of women death. The development of more sensitive diagnostic tests, which could present a faster response, lower cost, and could promote early diagnosis would increase the chances of survival. This study reports the development and optimization of an electrochemical aptasensor for the detection of HER2 protein, a breast cancer biomarker. Two sensing platforms were developed on gold screen-printed electrodes. The first platform is composed of self-assembled monolayer (SAM) made from mixture of thiolated DNA aptamers specific for HER2 and 1-mercapto-6-hexanol (MCH), while the second one is a ternary SAM composed of the same aptamer and 1,6-hexanethiol (HDT). Both platforms were further passivated with MCH and blocked with bovine serum albumin. The biosensors were characterized using electrochemical impedance spectroscopy to detect the target protein from 1 pg/mL to 1 µg/mL in phosphate buffered saline, diluted and undiluted human serum through charge transfer resistance value. The ternary SAM architecture shows a reduction of non-specific attachment to the electrode surface due to the HDT antifouling properties. In addition, this platform exhibits 172 pg/mL as limit of detection and a sensitivity of 4.12% per decade for undiluted serum compared with SAM architecture with the 179 pg/mL and 4.32% per decade, respectively. Electrochemical aptasensors are highly promising for medical diagnostic and ternary layers could improve the limit of detection.

Assuntos

RESUMO

In this work, a dual detection system based on an impedimetric immunosensor was developed for the first time for the simultaneous detection of anti-Trypanosoma cruzi and anti-Leishmania infantum antibodies in human and dog serum samples. The IBMP 8.1 and rLci1A/rLci2B recombinant antigens were immobilized over the surface of dual screen-printed carbon electrodes (W1 and W2) modified with poly (4-hydroxyphenylacetic acid). Under optimized conditions, the immunosensor recognized specific interactions for anti-T. cruzi antibodies up to a dilution of 1:10,240 and for anti-L. infantum up to 1:5120 in canine serum samples. Relative standard deviation (RSD) values of 2.8% for W1 and 3.6% for W2 were obtained for T. cruzi (W1) and L. infantum antigen (W2) samples in three different electrodes for 3 days (n = 9). The immunosensor was stored at 4 °C for 8 weeks, with activity retention of 70.2% in W1 and 78.2% in W2. The results using the recombinant proteins revealed that all antigens discriminated between negative and positive samples (p < 0.0001) in both dog and human groups, as well as no cross-reactivity could be detected among sera with other infections. With this approach, immunosensor-based diagnostic tests achieved 100% accuracy, suggesting that the antigens are eligible to enter Phase-II studies.

Assuntos

RESUMO

An electrochemical immunosensor was developed for the determination of apo-Tf (non-iron-bound) and holo-Tf (iron-bound) using polyclonal antibody transferrin (anti-Tf) immobilized at an electrode surface as a biorecognition platform. The monitoring was based on the anti-Tf binding with both Tf forms which allows the detection of cancer cells due to the constant iron cycle and the overexpression of anti-Tf on the cancer cell surface. The immunosensor characterization was performed using electrochemical impedance spectroscopy (EIS), which evaluated the impedimetric biorecognition of the antigens-antibody by the use of K4Fe(CN)6 redox group. The immunosensor was able to detect both forms of Tf in terms of charge transfer resistance (Rct). Analytical curves showed a limit of detection of 0.049 and 0.053 ng mL-1 for apo-Tf and holo-Tf, respectively. The immunosensor was applied to the detection of the two cancer cells A549 (lung carcinoma) and MCF-7 (breast carcinoma) and compared with BHK570, a healthy cell line. The impedimetric response of healthy cells differs significantly from that of the cancerous cells, as revealed by a Dunnett's test in 95% confidence level-ca. 102 cells mL-1-indicating the feasibility of the immunosensor to discriminate both types of cells. The indirect detection of anti-Tf based on apo-Tf and holo-Tf binding can be considered an advanced approach for cancer recognition. Graphical abstract.

Assuntos

RESUMO

A simple, cheap, and less aggressive immobilization procedure for biomolecules using reduced graphene oxide (rGO) was employed to prepare an impedimetric immunosensor for detection of staphylococcal enterotoxin A (SEA) from Staphylococcus aureus in milk samples. The scanning electron microscopy, cyclic voltammetry, and electrochemical impedance spectroscopy (EIS) were used to monitor the single steps of the electrode assembly process. The glassy carbon (GC)/rGO platform detected the antigen-antibody binding procedures of SEA with concentrations of 0.5 to 3.5 mg L-1 via impedance changes in a low frequency range. The impedimetric immunosensor was successfully applied for the determination of SEA in milk samples.

RESUMO

Zika virus (ZIKV) has recently become a global health challenge due to its rapid geographical expansion, since it is associated with serious neurological anomalies such as Guillain-Barré syndrome and microcephaly. Currently, the techniques for ZIKV diagnosis require labor-intensive, expensive and lengthy tests using sophisticated equipment. Moreover, false-positive or false-negative results can occur. In the present work, a DNA biosensor to detect ZIKV in real human serum samples was developed using an oxidized glassy carbon electrode (ox-GCE) modified with silsesquioxane-functionalized gold nanoparticles (AuNPs-SiPy). This nanohybrid was characterized by UV-Vis, FTIR and Raman spectroscopies, DLS, and XRD. The conditions for the immobilization of a ZIKV ssDNA probe on the electrode surface (ox-GCE-[AuNPs-SiPy]) were optimized by univariate and multivariate analysis. The optimized biosensor was characterized by CV, EIS and AFM experiments. The ZIKV target recognition was based on the variation of the charge transfer resistance (ΔRct) of the redox marker ([Fe(CN)6]3-/4-) used and the roughness (Rq) of the electrode surface. The proposed biosensor presented a LOD of 0.82 pmol L-1, with a linear range of 1.0 x10-12 - 1.0 x10-6 mol L-1. Moreover, the reported device showed a suitable stability and satisfactory sensitivity and selectivity to quantify ZIKV in human serum samples, which suggests its promising clinical applications for the early diagnosis of ZIKV-associated pathologies.

Assuntos

RESUMO

Pediatric adrenocortical carcinoma (pACC) is a rare and aggressive malignancy of high occurrence in Southern Brazil. pACC is characterized by the usual overproduction of dehydroepiandrosterone sulfate (DHEAS), whose detection in serum or plasma can be effective to the early diagnosis of the disease. Therefore, the present paper reports, for the first time, the construction and application of a label-free impedimetric immunosensor to detect DHEAS, which was based on the modification of an oxidized glassy carbon electrode with arginine-functionalized gold nanoparticles (AuNPs-ARG) and anti-DHEA IgM antibodies (ox-GCE/AuNPs-ARG/IgM). AuNPs-ARG was synthesized by a green route, and characterized by UV-VIS spectroscopy, FTIR, TEM, DLS, and XRD. The construction of ox-GCE/AuNPs-ARG/IgM was optimized through factorial design and response surface methodology. Cyclic voltammetry and electrochemical impedance spectroscopy measurements were employed to characterize the optimized immunosensor. The DHEAS detection principle was based on the variation of charge transfer resistance (∆Rct) relative to the Fe(CN)64-/3- electrochemical probe after immunoassays in the presence of the biomarker. A linear relationship between ∆Rct and DHEAS concentration was verified in the range from 10.0 to 110.0 µg dL-1, with a LOD of 7.4 µg dL-1. Besides the good sensitivity, the immunosensor displayed accuracy, stability, and specificity to detect DHEAS. The promising analytical performance of ox-GCE/AuNPs-ARG/IgM was confirmed by quantifying DHEAS in real patient plasma samples, with results that were comparable to the reference chemiluminescence assay. Our results suggest that the presented immunosensor can find clinical applications in the early diagnosis of pACC and to monitor DHEAS levels in other adrenal pathologies.

Assuntos