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Electrochemical techniques are commonly applied to micro total analysis system (μTAS) devices mainly due to its high sensitivity and miniaturization capacity. Among many electrochemical techniques, capacitively coupled contactless conductivity detection (C4 D) stands out for not requiring direct electrode-solution contact, avoiding several problems such as electrolysis, bubble formation, and metal degradation. Furthermore, the instrumentation required for C4 D measurements is compact, low cost, and easy to use, allowing in situ measurements to be performed even by nonspecialized personal. Contrarily, the production of metallic electrodes and microchannels adequate for C4 D measurements commonly requires specialized facilities and workers, increasing the costs of applying these methods. We propose alternatives to batch manufacture metallic electrodes and polymeric microchannels for C4 D analysis using more straightforward equipment and lower-cost materials. Three devices with different dielectric layer compositions and electrode sizes were tested and compared regarding their analytical performance. The constructed platforms have shown a reduction of more than 64% in cost when compared to traditional techniques and displayed good linearity (R2 ≥ 0.994), reproducibility (RSD ≤ 4.07%, n = 3), and limits of detection (≤0.26 mmol/L) when measuring standard NaCl samples. Therefore, the proposed methods were successfully validated and are available for further C4 D applications such as diagnosis of dry-eye syndrome.

This paper reports the development of a low-cost ( US$ 0.03 per device) immunosensor based on gold-modified screen-printed carbon electrodes (SPCEs). As a proof of concept, the immunosensor was tested for a fast and sensitive determination of S proteins from both SARS-CoV and SARS-CoV-2, by a single disposable device. Gold nanoparticles were electrochemically deposited via direct reduction of gold ions on the electrode using amperometry. Capture antibodies from spike (S) protein were covalently immobilized on carboxylic groups of self-assembled monolayers (SAM) of mercaptoacetic acid (MAA) attached to the gold nanoparticles. Label-free detection of S proteins from both SARS-CoV and SARS-CoV-2 was performed with electrochemical impedance spectroscopy (EIS). The immunosensor fabricated with 9 s gold deposition had a high performance in terms of selectivity, sensitivity, and low limit of detection (LOD) (3.16 pmol L

The 3D printing technology has gained ground due to its wide range of applicability. The development of new conductive filaments contributes significantly to the production of improved electrochemical devices. In this context, we report a simple method to producing an efficient conductive filament, containing graphite within the polymer matrix of PLA, and applied in conjunction with 3D printing technology to generate (bio)sensors without the need for surface activation. The proposed method for producing the conductive filament consists of four steps: (i) mixing graphite and PLA in a heated reflux system; (ii) recrystallization of the composite; (iii) drying and; (iv) extrusion. The produced filament was used for the manufacture of electrochemical 3D printed sensors. The filament and sensor were characterized by physicochemical techniques, such as SEM, TGA, Raman, FTIR as well as electrochemical techniques (EIS and CV). Finally, as a proof-of-concept, the fabricated 3D-printed sensor was applied for the determination of uric acid and dopamine in synthetic urine and used as a platform for the development of a biosensor for the detection of SARS-CoV-2. The developed sensors, without pre-treatment, provided linear ranges of 0.5-150.0 and 5.0-50.0 μmol L

Electrochemical techniques are commonly applied to micro total analysis system (μTAS) devices mainly due to its high sensitivity and miniaturization capacity. Among many electrochemical techniques, capacitively coupled contactless conductivity detection (C

We report on a wearable tear bioelectronic platform, integrating a microfluidic electrochemical detector into an eyeglasses nose-bridge pad, for non-invasive monitoring of key tear biomarkers. The alcohol-oxidase (AOx) biosensing fluidic system allowed real-time tear collection and direct alcohol measurements in stimulated tears, leading to the first wearable platform for tear alcohol monitoring. Placed outside the eye region this fully wearable tear-sensing platform addresses drawbacks of sensor systems involving direct contact with the eye as the contact lenses platform. Integrating the wireless electronic circuitry into the eyeglasses frame thus yielded a fully portable, convenient-to-use fashionable sensing device. The tear alcohol sensing concept was demonstrated for monitoring of alcohol intake in human subjects over multiple drinking courses, displaying good correlation to parallel BAC measurements. We also demonstrate for the first time the ability to monitor tear glucose outside the eye and the utility of wearable devices for monitoring vitamin nutrients in connection to enzymatic flow detector and rapid voltammetric scanning, respectively. These developments pave the way to build an effective eyeglasses system capable of chemical tear analysis.

We report on genosensors to detect an ssDNA sequence from the SARS-CoV-2 genome, which mimics the GU280 gp10 gene (coding the viral nucleocapsid phosphoprotein), using four distinct principles of detection and treating the data with information visualization and machine learning techniques. Genosensors were fabricated on either gold (Au) interdigitated electrodes for electrical and electrochemical measurements or on Au nanoparticles on a glass slide for optical measurements. They contained a matrix of 11-mercaptoundecanoic acid (11-MUA) self-assembled monolayer (SAM) onto which a layer of capture probe (cpDNA) sequence was immobilized. Detection was performed using electrical and electrochemical impedance spectroscopies and localized surface plasmon resonance (LSPR). The highest sensitivity was reached with impedance spectroscopy, including using a low-cost (US$ 100) homemade impedance analyzer. Complementary ssDNA sequences were detected with a detection limit of 0.5 aM (0.3 copy per µL). This performance may be attributed to the high sensitivity of the electrical impedance technique combined with an appropriate arrangement of the sequences on the electrodes and hybridization between the complementary sequences, as inferred from polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS). The selectivity of the genosensor was confirmed by plotting the impedance spectroscopy data with a multidimensional projection technique (interactive document mapping, IDMAP), where a clear separation was observed among the samples of the complementary DNA sequence at various concentrations and from buffer samples containing a non-complementary sequence and other DNA biomarkers. The diagnosis of SARS-CoV-2 mimicking sequences was also achieved with machine learning techniques applied to scanning electron microscope images taken from genosensors exposed to distinct concentrations of the complementary ssDNA sequences. In summary, the genosensors proposed here are promising for detecting SARS-CoV-2 genetic material (RNA) in biological fluids in point-of-care settings., The authors are thankful to CAPES (88887.510657/2020-00 and 88887.364257/2019-00), São Paulo Research Foundation (FAPESP) (2013/14262, 2016/01919-6, 2018/18953-8, 2018/19750-3, 2018/22214-6, 2019/00101-8, 2019/13514-9, 2016/23763-8, 2019/07811-0, 2020/02938-0), INEO, INCTBio grants (FAPESP 2014/50867-3) and CNPq (423952/2018-8, 465389/2014-7, and 401256/2020-0) for the financial support. The authors are grateful to Angelo Luiz Gobbi and Maria Helena de Oliveira Piazzetta (LMF/LNNano/CNPEM) for producing the Au electrodes.

Viruses are the causing agents for many relevant diseases, including influenza, Ebola, HIV/AIDS, and COVID-19. Its rapid replication and high transmissibility can lead to serious consequences not only to the individual but also to collective health, causing deep economic impacts. In this scenario, diagnosis tools are of significant importance, allowing the rapid, precise, and low-cost testing of a substantial number of individuals. Currently, PCR-based techniques are the gold standard for the diagnosis of viral diseases. Although these allow the diagnosis of different illnesses with high precision, they still present significant drawbacks. Their main disadvantages include long periods for obtaining results and the need for specialized professionals and equipment, requiring the tests to be performed in research centers. In this scenario, biosensors have been presented as promising alternatives for the rapid, precise, low-cost, and on-site diagnosis of viral diseases. This critical review article describes the advancements achieved in the last five years regarding electrochemical biosensors for the diagnosis of viral infections. First, genosensors and aptasensors for the detection of virus and the diagnosis of viral diseases are presented in detail regarding probe immobilization approaches, detection methods (label-free and sandwich), and amplification strategies. Following, immunosensors are highlighted, including many different construction strategies such as label-free, sandwich, competitive, and lateral-flow assays. Then, biosensors for the detection of viral-diseases-related biomarkers are presented and discussed, as well as point of care systems and their advantages when compared to traditional techniques. Last, the difficulties of commercializing electrochemical devices are critically discussed in conjunction with future trends such as lab-on-a-chip and flexible sensors.

To increase milk production, antibiotics are administered to animals to provide weight gain and to prevent or treat diseases. The inappropriate use of these substances can lead to antibiotic resistance and allergic reactions and toxic effects to milk consumers. We describe the development of a simple, fast, portable, and low-cost microfluidic paper-based analytical device (μPAD) to quantify sulfonamides in milk using the inhibition of the colorimetric reaction between carbonic anhydrase (CA) and 4-nitrophenyl acetate. The main advantages presented by the μPAD include reproducible batch production, simple application, and precise analysis without previous treatment. The µPAD displayed good linearity (R2 ≥ 0.986) in a wide range of sulfonamides in milk (2.5 to 40.0 µmol L−1), being selective for the drugs even in a highly complex matrix. We expect that this device allows in situ monitoring of milk quality, reducing the prejudicial conditions associated with high concentrations of sulfonamides in milk.

Nos últimos anos, o desenvolvimento de novas plataformas biossensoras trouxe importantes avanços à área médica, incluindo maior rapidez na detecção de biomarcadores, menor custo de análise e portabilidade. Essa tese traz a descrição do desenvolvimento de três tipos de biossensores que visam à auxiliar o diagnóstico de diferentes doenças. No primeiro capítulo, detalha-se a construção de um dispositivo microfluídico baseado em papel para a quantificação simultânea de dois biomarcadores da doença de Alzheimer: Fetuína B e Clusterina. O dispositivo foi capaz de quantificar as proteínas estudadas de maneira precisa e de baixo custo através do uso de técnicas eletroquímicas e colorimétricas, apresentando limites de detecção na ordem de p-nmol/L. O segundo capítulo trata do desenvolvimento de um imunossensor eletroquímico para a quantificação de melatonina. O dispositivo foi capaz de detectar a biomolécula em amostras biológicas rapidamente e com boa especificidade, diferenciando a melatonina de componentes com alta semelhança estrutural. Por fim, o terceiro capítulo descreve um genossensor eletroquímico para diagnóstico e determinação do traço da doença genética mais comum no Brasil, a anemia falciforme. O sensor foi capaz de diferenciar sequências normais de mutadas de maneira simples e rápida, sendo adequado para o diagnóstico e a determinação de seu traço. Esperamos que os dispositivos aqui apresentados levem à diagnósticos mais acessíveis fornecendo assim melhores condições de saúde à população. The development of new biosensor platforms has brought important advances to the medical field, including faster detection of biomarkers, lower cost of analysis and portability. This thesis provides a description of the development of three biosensors that aim to assist the diagnosis of three different diseases. The first chapter describes the construction of a paper-based microfluidic device for the simultaneous quantification of two Alzheimer\'s disease biomarkers: Fetuin B and Clusterin. The device was able to quantify the studied proteins in a precise and low cost manner through the use of electrochemical and colorimetric techniques, presenting limits of detection in the order of p-nmol/L. The second chapter details the the development of an electrochemical immunosensor for the quantification of melatonin. The device was able to detect the biomolecule in biological samples quickly and with good specificity, differentiating melatonin from components with high structural similarity. Finally, the third chapter describes an electrochemical genosensor for diagnosis and determination of the trait of the most common genetic disease in Brazil, sickle cell anemia. The sensor was able to differentiate normal and mutated sequences in a simple and fast manner, being suitable for the diseases diagnosis and the determination of its trait. Therefore, we expect the devices presented here to lead to more accessible diagnoses, providing better health conditions for the population.

Alzheimer's disease is a devastating condition characterized by a progressive and slow brain decay in elders. Here, we developed a paper-based lateral flow immunoassay for simultaneous and fast determination of Alzheimer's blood biomarkers, fetuin B and clusterin. Selective antibodies to targeted biomarkers were immobilized on gold nanoparticles (AuNPs) and deposited on paper pads. After adding the sample on the paper-based device, the biofluid laterally flows toward the selective antibody, permitting AuNP-Ab accumulation on the test zone, which causes a color change from white to pink. Image analysis was performed using a customized algorithm for the automatic recognition of the area of analysis and color clustering. Colorimetric detection was compared to electrochemical methods for the precise quantification of biomarkers. The best performance was found for the color parameter "L*". Good linearity (R2 = 0.988 and 0.998) and reproducibility (%RSD = 2.79% and 1.82%, N = 3) were demonstrated for the quantification of fetuin B and clusterin, respectively. Furthermore, the specificity of the immunosensor was tested on mixtures of proteins, showing negligible cross-reactivity and good performance in complex environments. We believe that our biosensor has a potential for early-stage diagnosis of Alzheimer's disease and toward a better understanding of Alzheimer's developing mechanisms.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by a progressive and irreversible cognitive decline. Currently, it affects 36 million people and due to population ageing it is estimated that in 2030 disease incidence will reach 60 million individuals. The precise diagnosis of AD is still a complex task, being mainly performed by cerebrospinal fluid (CSF) analysis or neuroimaging techniques such as positron emission tomography (PET) and magnetic resonance imaging (MRI). Despite being effective these techniques are expensive, time-consuming and not accessible for most part of the population. In this scenario biosensors are presented as promising alternatives for simple, rapid and low cost diagnosis of AD. In this revision we summarize the recent advances on biosensors that brings more accessibility to AD diagnosis. We introduce the most used biorecognition elements in miniaturized biosensing systems as well as AD biomarkers present in CSF, in plasma and in genetic material which can be used for disease identification even in early stages. The recent developed biosensors for AD diagnosis using optical, electrochemical and colorimetric techniques as well as their strategies and analytical performances are discussed. Advancements in signal amplification methodologies with nanomaterials to increase biosensors sensitivity are also presented. This review highlights the potential of biosensors to be used as an accurate and portable tool to improve the early AD diagnosis.

In this study, platinum electrodes were fabricated on the bio-based poly(ethylene terephthalate) (Bio-PET) substrates for the development of flexible electrochemical sensors for the detection of Parkinson's disease biomarkers. Dopamine was detected by voltammetric measurements, displaying a 3.5 × 10−5 mol L−1 to 8.0 × 10−4 mol L−1 linear range with a limit of detection of 5.1 × 10−6 mol L−1. Parkinson's disease protein 7 (PARK7/DJ-1) was successfully detected by electrochemical impedance spectroscopy after electrode functionalization with specific anti-PARK7/DJ-1 antibodies. In this case, analytical curves presented a linear behavior from 40 ng mL−1 to 150 ng mL−1 of PARK7/DJ-1 with a limit of detection of 7.5 ng mL−1. Besides, the electrodes did not suffer any change in the electrochemical response after manual tests of mechanical tension. The proposed sensor and immunosensor were applied for the determination of Parkinson's disease biomarkers concentrations found in the human body, being adequate as an alternative method to diagnose this disease.

Melatonin, the ‘chemical signal of darkness’, is responsible to regulate biological rhythms and different physiological processes. It is mainly produced by the pineal gland as a hormone in a rhythmic daily basis, but it may also be synthesized by other tissues, such as immune cells, under inflammatory conditions. Its abnormal circulating levels have been related to several diseases such as type 2 diabetes, Alzheimer’s disease and some types of cancer. Currently, melatonin is exclusively quantified by ELISA or radioimmunoassays, which although are very sensitive techniques and present low detection limits, usually require specialized personal and equipment, restricting the tests to a limited number of patients. To overcome such limitations, we developed a novel easy-to-use electrochemical immunosensor for rapid melatonin quantification. Anti-melatonin antibodies were immobilized into Indium tin oxide (ITO) platforms using (3-Aminopropyl)triethoxysilane (APTES), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) crosslinkers. The platforms were assayed with synthetic and biologically-present melatonin containing samples. The developed device displayed a linear response in the concentration range from 0.75 to 7.5 μmol/L and a limit of detection of 0.175 μmol/L using Electrochemical Impedance Spectroscopy (EIS) (R2 = 0.989) and 0.513 μmol/L using Cyclic Voltammetry (CV) (R² = 0.953) for synthetic melatonin. Furthermore, the sensors exhibited a good stability and reproducibility (3.45% and 2.87% for EIS and CV, respectively, n = 3), maintaining adequate response even after 30 days of assembly. On biologically-present melatonin-containing samples the device displayed a similar performance when compared to ELISA technique (deviation of 13.31%). We expect that the developed device contributes significantly to the medical area allowing precise and complete diagnosis of the diseases related to abnormal levels of melatonin.

Sickle cell anemia (SCA) is a common recessive genetic condition in which patients produce an abnormal form of hemoglobin. The disease is common mainly among African individuals and in parts of the continent up to 40 % of the population presents its genetic trait. Currently, disease diagnosis and trait determination are performed using polymerase chain reaction, liquid chromatography and electrophoresis. Although these methods present high sensitivity and are well established, they are costly and require specialized equipment to be performed. We developed an electrochemical genosensor for simple and low cost SCA trait determination. The device was based on the immobilization of single DNA strands containing the disease related mutation on gold platforms using the self-assembled monolayers technique. The determination of SCA trait was then performed using electrochemical impedance spectroscopy. The genosensor displayed a wide linear range (0.01 to 7.5 μmol L−1, R2=0.979), with a detection limit of 7.0 nmol L−1. Furthermore, the device was able to distinguish between DNA sequences containing or not the mutation (target and non-target sequences) with precision and great reproducibility (10.4 %, n=3). It is expected that such sensor increases the number of SCA trait determination, promoting early diagnosis and genetically counseling.

We present a review on the use of nanomaterials in point-of-care (POC) electrochemical biosensors applied to medicine, addressing their development and main advantages. We focused this chapter on the most relevant nanomaterials used in the nanomedicine field, which we understand to be metallic nanoparticles, carbon nanotubes, and graphene. The main strategies for nanomaterial synthesis and characterization are introduced and discussed, as well as the application of these materials in POC medical devices. Furthermore, the principal biological recognition elements and electrochemical techniques used in the development of biosensors are detailed. A final analysis and perspective on the subject is presented at the end of the chapter.

A obesidade tem aumentado dramaticamente nos últimos anos, tornando-se o maior fator de risco para o desenvolvimento de doenças cardiovasculares, vários tipos de câncer, e também do Diabetes mellitus tipo 2 (DM 2). A associação da obesidade com o desenvolvimento do DM 2 pode ser explicada, em parte, pela secreção alterada pelo tecido adiposo de adipocinas, como a adiponectina, um hormônio com propriedades antiinflamatórias e de sensibilização à insulina. O excesso do tecido adiposo regula negativamente a secreção de adiponectina, o que ocorre de 10 a 20 anos antes da hiperglicemia crônica, fazendo com que este hormônio seja um bom indicador para o diagnóstico preditivo de DM 2. A concentração de adiponectina em plasma é usualmente aferida por ELISA, método não amplamente utilizado devido ao seu alto preço, necessidade de pessoal qualificado e material e, dessa maneira, ainda praticamente inacessível à grande parcela da população. Sendo assim, o desenvolvimento de novas metodologias e de ferramentas de diagnóstico confiáveis e de baixo custo (que possam ser implementados pelo Sistema Único de Saúde SUS) é imprescindível. Aqui, desenvolvemos um biossensor simples e de baixo custo para detecção de adiponectina baseado na espectroscopia de impedância eletroquímica (EIS) ou voltametría cíclica (CV) usando plataformas nanoestruturadas contendo receptores transmembrana de adiponectina (AdipoR1 e AdipoR2) ou anticorpos anti-AdipoQ imobilizados em eletrodos de ouro. Os melhores resultados foram obtidos através do uso dos receptores AdipoR1/R2 em conjunto com CV. Neste caso, os biossensores foram capazes de detectar concentrações de adiponectina tão baixas quanto 7 nmol L-1 com uma faixa linear entre 0,01 a 0,75 mol L-1 de adiponectina, R²=0,992. O dispositivo apresentou ótima seletividade, estabilidade e reprodutibilidade (ca. 1,7% para n=3). Além disso, em amostras de plasma humano, o biossensor obteve resultados muito próximos aos obtidos pelo método pardão ELISA, com desvio de 14%. Esperamos que este estudo propicie maior acessibilidade ao diagnóstico preditivo do DM 2 através de dispositivos mais baratos, rápidos e portáteis e que um maior número indivíduos possam ser alertados e orientados, evitando o desenvolvimento posterior da doença. Obesity has increased dramatically in the last few years, becoming the biggest risk factor for development of cardiovascular diseases, several types of cancer and also the type 2 diabetes mellitus (DM 2). The association between obesity and DM 2 can be partially explained by the altered secretion of adiponectin hormone by the adipose tissue, which presents anti-inflammatory and insulin sensitizing properties. The excess of adipose tissue regulates negatively adiponectin secretion, which occurs 10 to 20 years before chronic hyperglycemia, making this hormone a great biomarker for predictive DM 2 diagnosis. Adiponectin plasma concentration is usually measured by ELISA, a method not widely used due to its high cost, personal and material demand, being, therefore, not accessible to part of the population. Therefore, the development of new diagnostic methodologies and tools trustable and low-cost (that can be implemented by Sistema Único de Saúde SUS) is crucial. Here we developed a simple and low-cost biosensor for adiponectin detection based on electrochemical impedance spectroscopy (EIS) or cyclic voltammetry (CV) using nanostructured platforms containing adiponectin transmembrane receptors (AdipoR1 and AdipoR2) or anti-AdipoQ antibodies immobilized on gold electrodes. The best results were achieved using AdipoR1/R2 receptors among with CV. In this case, biosensors were able to detect adiponectin at concentrations down to 7 nmol L-1 in a linear detection range from 0.01 to 0.75 mol L-1 of adiponectin, R²=0.992. The device displayed great selectivity, stability and reproducibility (ca. 1.7% for n=3). For human plasma samples, analyses using the biosensor and the ELISA technique presented similar results, with deviations of 14%. Therefore, we expect that this study leads to a greater accessibility to predictive DM 2 diagnoses through cheaper, faster and portable devices, so that a higher number of patients may be alerted and oriented, avoiding further disease development.

Sickle cell anemia (SCA) is a common recessive genetic condition in which patients produce hemoglobin S (HbS), an abnormal form of the protein, instead of hemoglobin A, the regular form. SCA is caused by the substitution of a single nitrogenous base adenine for thymine (GAG-> GTG), encoding valine instead of glutamine.1 Deoxygenation of HbS leads to its polymerization, damaging the cell membrane and reducing the lifetime of erythrocytes dramatically – which, in turn, leads to anemia. In some regions of Africa, up to 40% of the population possess its genetic trait, being considered a public health serious problem2. As the disease is possibly deadly, presents very low cure chances and has limited treatment, carriers determination is a key feature to assist in reproducibility decisions. Currently, trait determination is mainly performed by liquid chromatography, electrophoresis and polymerase chain reaction (PCR)3,4. These techniques are not widely used due to its complexity, lack of portability, high price and specialized personal and equipment demand. In this study we have developed an electrochemical genosensor for simple, low cost and rapid SCA carriers determination. Single DNA strands corresponding to the sequence of a SCA carrier were immobilized in gold platforms using the self-assembled-monolayers technique. The determination of SCA trait is then performed by analyzing the film’s charge transfer resistance using Electrochemical Impedance Spectroscopy. The genosensor was able to distinguish between healthy and unhealthy subjects with great precision besides displaying a wide linear range (0.01 to 7.5 µmol/L, R2 = 0.927). Furthermore, the device presented a detection limit of 7.0 nmol/L. We expect that such devices increases the number of SCA carriers determination, promoting early diagnosis and genetical counseling. References: 1 Barany, F. Genetic-Disease Detection and DNA Amplification Using Cloned Thermostable Ligase. Proceedings of the National Academy of Sciences of the United States of America 88, 189-193, doi:10.1073/pnas.88.1.189 (1991). 2 Diallo, D. & Tchernia, G. Sickle cell disease in Africa. Current Opinion in Hematology 9, 111-116, doi:10.1097/00062752-200203000-00005 (2002). 3 Ayatollahi, M. & Haghshenas, M. Application of the polymerase chain reaction to the diagnosis of sickle cell disease in Iran. Archive of Iranian Medicine 7, 84-88 (2004). 4 Bender, M. A. & Seibel, G. D. Sickle Cell Disease. (University of Washington, 2003).