Disposable immunosensor using a simple method for oriented antibody immobilization for label-free real-time detection of an oxidative stress biomarker implicated in cancer diseases

This work proposes a novel approach for a suitable orientation of antibodies (Ab) on an immunosensing platform, applied here to the determination of 8-hydroxy-2′-deoxyguanosine (8OHdG), a biomarker of oxidative stress that has been associated to chronic diseases, such as cancer. The anti-8OHdG was bound to an amine modified gold support through its Fc region after activation of its carboxylic functions. Non-oriented approaches of Ab binding to the platform were tested in parallel, in order to show that the presented methodology favored Ab/Ag affinity and immunodetection of the antigen. The immunosensor design was evaluated by quartz-crystal microbalance with dissipation, atomic force microscopy, electrochemical impedance spectroscopy (EIS) and square-wave voltammetry. EIS was also a suitable technique to follow the analytical behavior of the device against 8OHdG. The affinity binding between 8OHdG and the antibody immobilized in the gold modified platform increased the charge transfer resistance across the electrochemical set-up. The observed behavior was linear from 0.02 to 7.0 ng/mL of 8OHdG concentrations. The interference from glucose, urea and creatinine was found negligible. An attempt of application to synthetic samples was also successfully conducted. Overall, the presented approach enabled the production of suitably oriented Abs over a gold platform by means of a much simpler process than other oriented-Ab binding approaches described in the literature, as far as we know, and was successful in terms of analytical features and sample application.

Chitosan/AuNPs Modified Graphene Electrochemical Sensor for Label-Free Human Chorionic Gonadotropin Detection

A new immunosensor is presented for human chorionic gonadotropin (hCG), made by electrodepositing chitosan/gold-nanoparticles over graphene screen-printed electrode (SPE). The antibody was covalently bound to CS via its Fc-terminal. The assembly was controlled by electrochemical Impedance Spectroscopy (EIS) and followed by Fourier Transformed Infrared (FTIR). The hCG-immunosensor displayed linear response against the logarithm-hCG concentration for 0.1–25 ng/mL with limit of detection of 0.016 ng/mL. High selectivity was observed in blank urine and successful detection of hCG was also achieved in spiked samples of real urine from pregnant woman. The immunosensor showed good detection capability, simplicity of fabrication, low-cost, high sensitivity and selectivity.

Disposable immunosensor with simple antibody orientation for label-free real-time detection of a cancer biomarker

This work proposes a novel approach for a suitable orientation of antibodies (Ab) on an immunosensing platform, applied here to the determination of 8-hydroxy-2’-deoxyguanosine (8OHdG), a biomarker of oxidative stress that has been associated to chronic diseases, such as cancer. The Anti-8OHdG was bound to an amine modified gold support through its Fc region after activation of its carboxylic functions. Non-oriented approaches of Ab binding to the platform were tested in parallel, in order to show that the presented proposal favored Ab/Ag affinity. The immunosensor design was evaluated by Quartz-Crystal microbalance with Dissipation, Atomic Force Microscopy, Electrochemical Impedance Spectroscopy (EIS) and Square-Wave Voltammetry. EIS was also a suitable technique to follow the analytical behavior of the device against 8OHdG. The affinity binding between 8OHdG and the antibody immobilized in the gold modified platform increased the charged transfer resistance across the electrochemical sep-up. The observed behavior was linear from 0.02 to 7.0 ng/mL of 8OHdG concentrations. The interference from Glucose, Urea and Creatinine was found negligible. An attempt of application to synthetic samples was also successfully conducted. Overall, the presented approach enabled the production of suitably oriented Abs over a gold platform by means of a much simpler process than other oriented-Ab binding approaches described in the literature, as far as we know, and was successful in terms of analytical features and sample application.

Label-free human chorionic gonadotropin detection at picogram levels using oriented antibodies bound to graphene screen-printed electrodes

Human chorionic gonadotropin (hCG) is a key diagnostic marker of pregnancy and an important biomarker for cancers in the prostate, ovaries and bladder and therefore of great importance in diagnosis. For this purpose, a new immunosensor of screen-printed electrodes (SPEs) is presented here. The device was fabricated by introducing a polyaniline (PANI) conductive layer, via in situ electropolymerization of aniline, onto a screen-printed graphene support. The PANI-coated graphene acts as the working electrode of a three terminal electrochemical sensor. The working electrode is functionalised with anti-hCG, by means of a simple process that enabled oriented antibody binding to the PANI layer. The antibody was attached to PANI following activation of the –COOH group at the Fc terminal. Functionalisation of the electrode was analysed and optimized using Electrochemical Impedance Spectroscopy (EIS). Chemical modification of the surface was characterised using Fourier transform infrared, and Raman spectroscopy with confocal microscopy. The graphene–SPE–PANI devices displayed linear responses to hCG in EIS assays from 0.001 to 50 ng mL−1 in real urine, with a detection limit of 0.286 pg mL−1. High selectivity was observed with respect to the presence of the constituent components of urine (urea, creatinine, magnesium chloride, calcium chloride, sodium dihydrogen phosphate, ammonium chloride, potassium sulphate and sodium chloride) at their normal levels, with a negligible sensor response to these chemicals. Successful detection of hCG was also achieved in spiked samples of real urine from a pregnant woman. The immunosensor developed is a promising tool for point-of-care detection of hCG, due to its excellent detection capability, simplicity of fabrication, low-cost, high sensitivity and selectivity.

Specific label-free and real-time detection of oxidized low density lipoprotein (oxLDL) using an immunosensor with three monoclonal antibodies

Increased levels of plasma oxLDL, which is the oxidized fraction of Low Density Lipoprotein (LDL), are associated with atherosclerosis, an inflammatory disease, and the subsequent development of severe cardiovascular diseases that are today a major cause of death in modern countries. It is therefore important to find a reliable and fast assay to determine oxLDL in serum. A new immunosensor employing three monoclonal antibodies (mAbs) against oxLDL is proposed in this work as a quick and effective way to monitor oxLDL. The oxLDL was first employed to produce anti-oxLDL monoclonal antibodies by hybridoma cells that were previously obtained. The immunosensor was set-up by selfassembling cysteamine (Cyst) on a gold (Au) layer (4 mm diameter) of a disposable screen-printed electrode. Three mAbs were allowed to react with N-hydroxysuccinimide (NHS) and ethyl(dimethylaminopropyl)carbodiimide (EDAC), and subsequently incubated in the Au/Cys. Albumin from bovine serum (BSA) was immobilized further to ensure that other molecules apart from oxLDL could not bind to the electrode surface. All steps were followed by various characterization techniques such as electrochemical impedance spectroscopy (EIS) and square wave voltammetry (SWV). The analytical operation of the immunosensor was obtained by incubating the sensing layer of the device in oxLDL for 15 minutes, prior to EIS and SWV. This was done by using standard oxLDL solutions prepared in foetal calf serum, in order to simulate patient's plasma with circulating oxLDL. A sensitive response was observed from 0.5 to 18.0 mg mL 1 . The device was successfully applied to determine the oxLDL fraction in real serum, without prior dilution or necessary chemical treatment. The use of multiple monoclonal antibodies on a biosensing platform seemed to be a successful approach to produce a specific response towards a complex multi-analyte target, correlating well with the level of oxLDL within atherosclerosis disease, in a simple, fast and cheap way.

A label-free DNA aptamer-based impedance biosensor for the detection of E. coli outer membrane proteins

A label-free DNA aptamer-based impedance biosensor for the detection of E. coli outer membrane proteins (OMPs) was developed. Two single stranded DNA sequences were tested as recognition elements and compared. The aptamer capture probes were immobilized, with and without 6-mercapto-1-hexanol (MCH) on a gold electrode. Each step of the modification process was characterized by Faradaic impedance spectroscopy (FIS). A linear relationship between the electron-transfer resistance (Ret) and E. coli OMPs concentration was demonstrated in a dynamic detection range of 1 × 10−7–2 × 10−6 M. Moreover, the aptasensor showed selectivity despite the presence of other possible water contaminates and could be regenerated under low pH condition. The developed biosensor shows great potential to be incorporated in a biochip and used for in situ detection of E. coli OMPs in water samples.

Label-free Detection of Microcystin-LR in Waters Using Real-Time Potentiometric Biosensors Based on Single-Walled Carbon Nanotubes Imprinted Polymers

Microcystin-LR (MC-LR) is a dangerous toxin found in environmental waters, quantified by high performance liquid chromatography and/or enzyme-linked immunosorbent assays. Quick, low cost and on-site analysis is thus required to ensure human safety and wide screening programs. This work proposes label-free potentiometric sensors made of solid-contact electrodes coated with a surface imprinted polymer on the surface of Multi-Walled Carbon NanoTubes (CNTs) incorporated in a polyvinyl chloride membrane. The imprinting effect was checked by using non-imprinted materials. The MC-LR sensitive sensors were evaluated, characterized and applied successfully in spiked environmental waters. The presented method offered the advantages of low cost, portability, easy operation and suitability for adaptation to flow methods.

Isoflavones in food supplements: chemical profile, label accordance and permeability study in Caco-2 cells

Consumers nowadays are playing an active role in their health-care. A special case is the increasing number of women, who are reluctant to use exogenous hormone therapy for the treatment of menopausal symptoms and are looking for complementary therapies. However, food supplements are not clearly regulated in Europe. The EFSA has only recently begun to address the issues of botanical safety and purity regulation, leading to a variability of content, standardization, dosage, and purity of available products. In this study, isoflavones (puerarin, daidzin, genistin, daidzein, glycitein, genistein, formononetin, prunetin, and biochanin A) from food supplements (n = 15) for menopausal symptoms relief are evaluated and compared with the labelled information. Only four supplements complied with the recommendations made by the EC on the tolerable thresholds. The intestinal bioavailability of these compounds was investigated using Caco-2 cells. The apparent permeability coefficients of the selected isoflavonoids across the Caco-2 cells were affected by the isoflavone concentration and product matrix.

Teas, dietary supplements and fruit juices: a comparative study regarding antioxidant activity and bioactive compounds

Nowadays, new emerging products claiming antioxidant properties are becoming more frequent. However, information about this topic in their labels is usually scarce. In this paper, we analyzed total phenolics, total flavonoids and ascorbic acid contents, as well as DPPH scavenging activity of several commercial samples, namely green tea and other herbal infusions, dietary supplements, and fruit juices, available in the Portuguese market. In general, beverages containing green tea and hibiscus showed higher phenolics contents (including flavonoids) and antioxidant activity than those without these ingredients. A borututu infusion presented the lowest concentrations of bioactive compounds and scavenging activity, due to the low recommended amount of plant to prepare the beverage. Some juices without antioxidant claims in the label presented similar values to those with it.

Devolução de doses unitárias: análise de uma prática diária

A Distribuição Individual Diária em Dose Unitária deve assegurar o maior rigor no Processo de Distribuição de Medicamentos, o que nem sempre se verificou no Hospital Pedro Hispano, dado que as Doses Unitárias devolvidas pelos Serviços de Internamento, nem sempre seguiam os Parâmetros de Qualidade. Assim, desenvolveu-se um estudo quasi-experimental, longitudinal, prospetivo e analítico, entre 1 de Março a 30 de Junho de 2012, no qual se analisaram as Doses Unitárias, devolvidas pelos Serviços de Internamento da Unidade Local de Saúde de Matosinhos a funcionam com Distribuição Individual Diária em Dose Unitária, tendo-se recolhido as não conformes relativamente aos Parâmetros de Qualidade instituídos. Na 1.ª fase do estudo (Recolha de amostras), foram contabilizadas 337 amostras, correspondentes a uma perda de 471 €. Na 2.ª fase do estudo (Implementação de acções correctivas às amostras previamente recolhidas), foram rejeitadas definitivamente 129 amostras, correspondentes a uma perda de 198 €. Na 4.ª fase do estudo (Segunda recolha de amostras), foram contabilizadas 228 amostras, correspondentes a uma perda de 199 €. A 3.ª fase do estudo incluiu a divulgação dos resultados às enfermeiras chefes dos serviços envolvidos no estudo e a 5.ª fase a comparação dos resultados obtidos na 1.ª, 2.ª e 4.ª fase do estudo. As amostras foram recolhidas em 16 serviços de internamento, sendo os valores monetários mais relevantes associados ao Espessante para alimentos líquidos e ao Imipeno + Cilastatina. A prática observada e sujeita a estudo aumenta os riscos associados ao consumo do medicamento e os custos relativos ao Processo, sendo de eliminar. A sua monitorização deve constituir uma rotina, uma vez que houve mudança de comportamentos. A redução/eliminação desta prática não conforme conduz à integração de novas tarefas com valor acrescentado, com aumento da fiabilidade do Processo.

Analysing TDMA with slot skipping

We propose a schedulability analysis for a particular class of time division multiple access (TDMA) networks, which we label as TDMA/SS. SS stands for slot skipping, reflecting the fact that a slot is skipped whenever it is not used. Hence, the next slot can start earlier in benefit of hard real-time traffic. In the proposed schedulability analysis, we assume knowledge of all message streams in the system, and that each node schedules messages in its output queue according to a rate monotonic policy (as an example). We present the analysis in two steps. Firstly, we address the case where a node is only permitted to transmit a maximum of one message per TDMA cycle. Secondly, we generalise the analysis to the case where a node is assigned a budget of messages per TDMA cycle it may transmit. A simple algorithm to assign budgets to nodes is also presented.

Marcação CE de dispositivos médicos e segurança em software

Com o crescente aumento da Teleradiologia, sentiu-se necessidade de criar mais e melhores softwares para sustentar esse crescimento. O presente trabalho pretende abordar a temática da certificação de software e a sua marcação CE, pois para dar entrada no mercado Europeu todos os Dispositivos Médicos (DM) têm de estar devidamente certificados. Para efetuar a marcação CE e a certificação serão estudadas normas e normativos adequados para marcação de DM ao nível Europeu e também dos Estados Unidos da América. A temática da segurança de dados pessoais será também estudada de forma a assegurar que o dispositivo respeite a legislação em vigor. Este estudo tem como finalidade a certificação de um software proprietário da efficientia sysPACS, um serviço online abrangente, que permite a gestão integrada do armazenamento e distribuição de imagens médicas para apoio ao diagnóstico.

Avaliação do desempenho energético de um grande edifício de serviços existente

Nesta dissertação pretende-se caracterizar o desempenho energético de um grande edifício de serviços existente, da tipologia ensino, avaliar e identificar potenciais medidas que melhorem aquele desempenho, permitindo, em complemento, determinar a sua classificação energética no âmbito da legislação vigente. A pertinência do estudo prende-se com a avaliação do desempenho energético dos edifícios e com o estudo de medidas de melhoria que permitam incrementar a eficiência energética, por recurso a um programa de simulação energética dinâmica certificado – DesignBuilder e tendo em conta a regulamentação portuguesa em vigor. Inicialmente procedeu-se à modelação do edifício com recurso ao programa DesignBuilder, e, simultaneamente, realizou-se um levantamento de todas as suas características ao nível de geometria, pormenores construtivos, sistemas AVAC e de iluminação e fontes de energia utilizadas. Com vista à caracterização do modo de operação do edifício, foi realizado um levantamento dos perfis reais de utilização em termos de ocupação, iluminação e equipamentos para os vários espaços. Foram realizadas medições de caudais de ar novo e da temperatura do ar, em alguns equipamentos e alguns espaços específicos. Foram realizadas medições em tempo real e leituras de contagens da energia eléctrica utilizada, quer em período de aulas quer em período de férias, que permitiram a desagregação das facturas da energia eléctrica que se apresentam globais para o campus do ISEP. Foram realizadas leituras de contagens de gás natural. Em sequência, foi realizada a simulação energética dinâmica com o intuito de ajustar o modelo criado aos consumos reais e de analisar medidas de melhoria que lhe conferissem um melhor desempenho energético. Essas medidas são agrupadas em quatro tipos: - Medidas de natureza comportamental; - Medidas de melhoria da eficiência energética nos sistemas de iluminação; - Medidas de melhoria de eficiência energética nos sistemas AVAC;- Medidas que visam a introdução de energias de fonte renovável; Em sequência, foi elaborada a simulação nominal e calculados os indicadores de eficiência energética com vista à respectiva classificação energética do edifício, tendo o edifício apresentado uma Classe Energética D de acordo com a escala do SCE. Finalmente, foi avaliado o impacto das diferentes medidas de melhoria identificadas e com potencial de aplicação, isto é, que apresentaram um retorno simples do investimento inferior a oito anos, tanto ao nível do desempenho energético real do edifício, como ao nível da sua classificação energética. De onde se concluiu que existe um potencial de 7% de redução nos consumos energéticos actuais do edifício e de 18% se o funcionamento do edifício for em pleno, ou seja, se todos os seus sistemas estiverem efectivamente em funcionamento, e que terá impacto na classificação energética alcançado uma Classe Energética C.

Desenvolvimento de código pictográfico para identificação de medicamentos

Trabalho de Projeto

Molecule: Sistema de organização e visualização de tags

No dia-a-dia existe regularmente a necessidade de rotular um item com informação adicional de forma a poder ser mais facilmente recuperado ou identificado posteriormente. Diversas plataformas permitem que os utilizadores rotulem recursos com tags que habitualmente são partilhadas com outros utilizadores. Assim, ao longo do tempo foram propostas várias formas de visualização das tags associados aos recursos, com o intuito de não só facilitar aos utilizadores a pesquisa dos mesmos, mas também permitir a visualização do tag space. A nuvem de tags destaca-se como a forma mais comum de visualização. Este documento apresenta um estudo efetuado sobre formas de visualização de tags, as suas vantagens e limitações, e propõe uma forma de visualização alternativa. Sugere-se também uma nova interpretação sobre como pesquisar e visualizar recursos com tags associadas: o sistema Molecule, uma solução viável e inovadora, para vários dos problemas associados à tradicional nuvem de tags que, para além de permitir aos seus utilizadores associem tags aos s recursos, proporciona uma abordagem multivista para os mesmos navegarem no tag space e pesquisarem informação.