Caracterização material e conservação e restauro de um painel de azulejos do séc. XVII do Ecomuseu do Seixal, Portugal

Dissertação para obtenção do Grau de Mestre em Conservação e Restauro, Especialização em Cerâmica e Vidro

Haemoglobin smart plastic antibody material tailored with charged binding sites on silica nanoparticles: its application as an ionophore in potentiometric transduction

This work uses surface imprinting to design a novel smart plastic antibodymaterial (SPAM) for Haemoglobin (Hb). Charged binding sites are described here for the first time to tailor plastic antibody nanostructures for a large size protein such as Hb. Its application to design small, portable and low cost potentiometric devices is presented. The SPAM material was obtained by linking Hb to silica nanoparticles and allowing its ionic interaction with charged vinyl monomers. A neutral polymeric matrix was created around these and the imprinted protein removed. Additional materials were designed in parallel acting as a control: a neutral imprinted material (NSPAM), obtained by removing the charged monomers from the procedure, and the Non-Imprinted (NI) versions of SPAM and NSPAM by removing the template. SEM analysis confirmed the surface modification of the silica nanoparticles. All materials were mixed with PVC/plasticizer and applied as selective membranes in potentiometric transduction. Electromotive force (emf) variations were detected only for selective membranes having a lipophilic anionic additive in the membrane. The presence of Hb inside these membranes was evident and confirmed by FTIR, optical microscopy and Raman spectroscopy. The best performance was found for SPAM-based selective membranes with an anionic lipophilic additive, at pH 5. The limits of detection were 43.8 mg mL 1 and linear responses were obtained down to 83.8 mg mL 1, with an average cationic slope of +40 mV per decade. Good selectivity was also observed against other coexisting biomolecules. The analytical application was conducted successfully, showing accurate and precise results.

Oriented Tailoring of Plastic Antibodies for Prostate Specific Antigen and Application of the Imprinted Material as Ionophore in Potentiometric Detection

Prostate Specific Antigen (PSA) is the biomarker of choice for screening prostate cancer throughout the population, with PSA values above 10 ng/mL pointing out a high probability of associated cancer1. According to the most recent World Health Organization (WHO) data, prostate cancer is the commonest form of cancer in men in Europe2. Early detection of prostate cancer is thus very important and is currently made by screening PSA in men over 45 years old, combined with other alterations in serum and urine parameters. PSA is a glycoprotein with a molecular mass of approximately 32 kDa consisting of one polypeptide chain, which is produced by the secretory epithelium of human prostate. Currently, the standard methods available for PSA screening are immunoassays like Enzyme-Linked Immunoabsorbent Assay (ELISA). These methods are highly sensitive and specific for the detection of PSA, but they require expensive laboratory facilities and high qualify personal resources. Other highly sensitive and specific methods for the detection of PSA have also become available and are in its majority immunobiosensors1,3-5, relying on antibodies. Less expensive methods producing quicker responses are thus needed, which may be achieved by synthesizing artificial antibodies by means of molecular imprinting techniques. These should also be coupled to simple and low cost devices, such as those of the potentiometric kind, one approach that has been proven successful6. Potentiometric sensors offer the advantage of selectivity and portability for use in point-of-care and have been widely recognized as potential analytical tools in this field. The inherent method is simple, precise, accurate and inexpensive regarding reagent consumption and equipment involved. Thus, this work proposes a new plastic antibody for PSA, designed over the surface of graphene layers extracted from graphite. Charged monomers were used to enable an oriented tailoring of the PSA rebinding sites. Uncharged monomers were used as control. These materials were used as ionophores in conventional solid-contact graphite electrodes. The obtained results showed that the imprinted materials displayed a selective response to PSA. The electrodes with charged monomers showed a more stable and sensitive response, with an average slope of -44.2 mV/decade and a detection limit of 5.8X10-11 mol/L (2 ng/mL). The corresponding non-imprinted sensors showed smaller sensitivity, with average slopes of -24.8 mV/decade. The best sensors were successfully applied to the analysis of serum samples, with percentage recoveries of 106.5% and relatives errors of 6.5%.

Electrochemical biosensor based on biomimetic material for myoglobin detection

A novel reusable molecularly imprinted polymer (MIP) assembled on a polymeric layer of carboxylated poly(vinyl chloride) (PVCsingle bondCOOH) for myoglobin (Myo) detection was developed. This polymer was casted on the gold working area of a screen printed electrode (Au-SPE), creating a novel disposable device relying on plastic antibodies. Electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and Fourier transform infrared spectroscopy (FTIR) studies confirmed the surface modification. The MIP/Au-SPE devices displayed a linear behaviour in EIS from 0.852 to 4.26 μg mL−1, of positive slope 6.50 ± 1.48 (kΩ mL μg−1). The limit of detection was 2.25 μg mL−1. Square wave voltammetric (SWV) assays were made in parallel and showed linear responses between 1.1 and 2.98 μg mL−1. A current decrease was observed against Myo concentration, producing average slopes of −0.28 ± 0.038 μA mL μg−1. MIP/Au-SPE also showed good results in terms of selectivity. The error% found for each interfering species were 7% for troponin T (TnT), 11% for bovine serum albumin (BSA) and 2% for creatine kinase MB (CKMB), respectively. Overall, the technical modification over the Au-SPE was found a suitable approach for screening Myo in biological fluids.

Smart Plastic Antibody Material (SPAM) tailored on disposable screen printed electrodes for protein recognition: application to Myoglobin detection

This work introduces two major changes to the conventional protocol for designing plastic antibodies: (i) the imprinted sites were created with charged monomers while the surrounding environment was tailored using neutral material; and (ii) the protein was removed from its imprinted site by means of a protease, aiming at preserving the polymeric network of the plastic antibody. To our knowledge, these approaches were never presented before and the resulting material was named here as smart plastic antibody material (SPAM). As proof of concept, SPAM was tailored on top of disposable gold-screen printed electrodes (Au-SPE), following a bottom-up approach, for targeting myoglobin (Myo) in a point-of-care context. The existence of imprinted sites was checked by comparing a SPAM modified surface to a negative control, consisting of similar material where the template was omitted from the procedure and called non-imprinted materials (NIMs). All stages of the creation of the SPAM and NIM on the Au layer were followed by both electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). AFM imaging was also performed to characterize the topography of the surface. There are two major reasons supporting the fact that plastic antibodies were effectively designed by the above approach: (i) they were visualized for the first time by AFM, being present only in the SPAM network; and (ii) only the SPAM material was able to rebind to the target protein and produce a linear electrical response against EIS and square wave voltammetry (SWV) assays, with NIMs showing a similar-to-random behavior. The SPAM/Au-SPE devices displayed linear responses to Myo in EIS and SWV assays down to 3.5 μg/mL and 0.58 μg/mL, respectively, with detection limits of 1.5 and 0.28 μg/mL. SPAM materials also showed negligible interference from troponin T (TnT), bovine serum albumin (BSA) and urea under SWV assays, showing promising results for point-of-care applications when applied to spiked biological fluids.

Synthesis of an antibody-like material for the detection of Albumin

6th Graduate Student Symposium on Molecular Imprinting

Smart Plastic Antibody Material for Hemoglobin Tailored by Silica Surface Imprinting and with Charged Binding Sites: Its use as Ionophore in Potentiometric Transduction

JORNADAS DE ELECTROQUÍMICA E INOVAÇÃO 2013

A novel antibody-like material for breast cancer antigen CA15-3, used to track breast cancer by potentiometric transduction

This work presents the development of a low cost sensor device for the diagnosis of breast cancer in point-of-care, made with new synthetic biomimetic materials inside plasticized poly(vinyl chloride), PVC, membranes, for subsequent potentiometric detection. This concept was applied to target a conventional biomarker in breast cancer: Breast Cancer Antigen (CA15-3). The new biomimetic material was obtained by molecularly-imprinted technology. In this, a plastic antibody was obtained by polymerizing around the biomarker that acted as an obstacle to the growth of the polymeric matrix. The imprinted polymer was specifically synthetized by electropolymerization on an FTO conductive glass, by using cyclic voltammetry, including 40 cycles within -0.2 and 1.0 V. The reaction used for the polymerization included monomer (pyrrol, 5.0×10-3 mol/L) and protein (CA15-3, 100U/mL), all prepared in phosphate buffer saline (PBS), with a pH of 7.2 and 1% of ethylene glycol. The biomarker was removed from the imprinted sites by proteolytic action of proteinase K. The biomimetic material was employed in the construction of potentiometric sensors and tested with regard to its affinity and selectivity for binding CA15-3, by checking the analytical performance of the obtained electrodes. For this purpose, the biomimetic material was dispersed in plasticized PVC membranes, including or not a lipophilic ionic additive, and applied on a solid conductive support of graphite. The analytical behaviour was evaluated in buffer and in synthetic serum, with regard to linear range, limit of detection, repeatability, and reproducibility. This antibody-like material was tested in synthetic serum, and good results were obtained. The best devices were able to detect 5 times less CA15-3 than that required in clinical use. Selectivity assays were also performed, showing that the various serum components did not interfere with this biomarker. Overall, the potentiometric-based methods showed several advantages compared to other methods reported in the literature. The analytical process was simple, providing fast responses for a reduced amount of analyte, with low cost and feasible miniaturization. It also allowed the detection of a wide range of concentrations, diminishing the required efforts in previous sample pre-treating stages.

Sol-gel biomimetic material designed to target CEA cancer biomarker

1st ASPIC International Congress

Cravo de Pascal Taskin (1782): caracterização material da policromia exterior

Dissertação para obtenção do Grau de Mestre em Conservação e Restauro, Especialização em Ciências da Conservação

A Visual Programming Framework for Wireless Sensor Networks in Smart Home Applications

International Conference on Intelligent Sensors, Sensor Networks and Information Processing (ISSNIP 2015). 7 to 9, Apr, 2015. Singapure, Singapore.

Autonomous Surface Vehicle Docking Manoeuvre with Visual Information

This work presents a hybrid coordinated manoeuvre for docking an autonomous surface vehicle with an autonomous underwater vehicle. The control manoeuvre uses visual information to estimate the AUV relative position and attitude in relation to the ASV and steers the ASV in order to dock with the AUV. The AUV is assumed to be at surface with only a small fraction of its volume visible. The system implemented in the autonomous surface vehicle ROAZ, developed by LSA-ISEP to perform missions in river environment, test autonomous AUV docking capabilities and multiple AUV/ASV coordinated missions is presented. Information from a low cost embedded robotics vision system (LSAVision), along with inertial navigation sensors is fused in an extended Kalman filter and used to determine AUV relative position and orientation to the surface vehicle The real time vision processing system is described and results are presented in operational scenario.

O treino da perceção visual e da consciência fonológica como promotor da fluência da leitura num aluno do 2º ciclo do ensino básico

Aprender a ler é um dos maiores desafios que as crianças enfrentam quando entram para a escola. A dificuldade no domínio do código alfabético, nos níveis da consciência fonológica e a falta de fluência na leitura são fatores que interferem em larga escala na aprendizagem global dos alunos. Habilitar um aluno para a prática da leitura é um estímulo que tem vindo a dar origem a várias investigações e intervenções no campo da educação. Este projeto descreve dois programas de treino: “Programa de treino da percepção Visual” e “Programa de promoção do desenvolvimento da consciência fonológica”, num aluno do 2º ciclo do ensino básico com dificuldade de fluência na leitura, ao longo de quinze aulas de 90 minutos. No que respeita aos resultados do primeiro estudo, que teve por base o “Programa de treino da percepção visual”, não foram encontradas diferenças relevantes quanto ao seu efeito na fluência da leitura do aluno. No entanto, no segundo estudo, que se centrou na aplicação do “Programa de promoção do desenvolvimento da consciência fonológica” em complemento com o “Programa de treino da percepção visual”, mostrou que o aluno ficou mais fluente na leitura diminuindo o número de erros de precisão (substituições, omissões, inversões, adições e erros complexos). Assim, sugere-se uma monotorização sistemática das aprendizagens dos alunos para que as intervenções possam ser cada vez mais precoces e direcionadas para as suas necessidades.

Otosclerose - Correlação Clínica e Imagiológica

Objectivos: Identificar factores que influenciem os resultados no pós-operatório em otosclerose, nomeadamente a tomografia computorizada (TC). Material e métodos: Estudo retrospectivo de 42 doentes. Análise estatística com o teste de T Student e o estudo de Correlações de Pearson. Resultados: O Gap Aero-Ósseo (GAO) médio no pré e pós operatório foi, respectivamente, 42 dB e 13 dB. Das 11 TC avaliadas, verificaram-se focos activos em 4 ouvidos e inactivos em 5. Uma das TC foi considerada normal. Houve um predomínio de focos do grupo 1 (7 ouvidos). Não houve diferença estatisticamente significativa do ganho da audição quanto à presença de entalhe de Carhart e ao GAO pré operatório (p>0.05). Embora houvesse uma correlação positiva entre a actividade do foco encontrada na TC e o ganho audiométrico, esta relação não foi estatisticamente significativa (p=0.094). Conclusão: A TC é um meio de imagem que pode auxiliar as decisões terapêuticas da otosclerose.