Sol-Gel Chemistry in Biosensing Devices of Electrical Transduction: Application to CEA Cancer Biomarker

Sol-gel chemistry allows the immobilization of organic molecules of biological origin on suibtable solid supports, permitting their integration into biosensing devices widening the possibility of local applications. The present work is an application of this principle, where the link between electrical receptor platform and the antibody acting as biorecognition element is made by sol-gel chemistry. The immunosensor design was targeted for carcinoembryonic antigen (CEA), an important biomarker for screening the colorectal cancer, by electrochemical techniques, namely electrochemical impedance spectroscopy (EIS) and square wave voltammetry (SVW). The device displayed linear behavior to CEA in EIS and in SWV assays ranging from 0.50 to 1.5ng/mL, and 0.25 to 1.5ng/mL, respectively. The corresponding detection limits were 0.42 and 0.043 ng/mL. Raman spectroscopy was used to characterize the surface modifications on the conductive platform (FTO glass). Overall, simple sol-gel chemistry was effective at the biosensing design and the presented approach can be a potential method for screening CEA in point-of-care, due to the simplicity of fabrication, short response time and low cost. - See more at: http://www.eurekaselect.com/127192/article#sthash.m1AWhINx.dpuf

Sol-gel biomimetic material designed to target CEA cancer biomarker

1st ASPIC International Congress

Digit Ratio (2D:4D) in Newborns: Influences of Prenatal Testosterone and Maternal Environment

Introduction: The 2D:4D digit ratio is sexually-dimorphic, probably due to testosterone action through the perinatal period. We characterize the 2D:4D ratio in newborn (NB) infants, in between the pre- and postnatal surges of testosterone, and relate it to the mother's 2D:4D and to testosterone levels in the amniotic fluid (AF). Subjects and methods: Testosterone was assayed in samples of maternal plasma and AF collected at amniocentesis. Shortly after birth, 106 NBs and their mothers were measured for 2D:4D ratio. Results: NB males had lower mean 2D:4D ratios than females but this dimorphism was significant only for the left hand (males: 0.927; females: 0.950; p=0.004). Mothers who had sons had lower 2D:4D ratios than those who had daughters and the mother's 2D:4D were higher than those of NBs regardless of sex. Both hands of NB females were negatively correlated with AF testosterone and positively correlated with the mother's 2D:4D, but males showed no significant associations. Maternal plasma testosterone also showed a negative weak correlation with NB's digit ratio in both sexes. Conclusions: Sexual dimorphism at birth was only significant for the left hand, in contrast with reports of greater right hand dimorphism, suggesting that postnatal testosterone is determinant for 2D:4D stabilization. The lower 2D:4D ratios in mothers who had sons support claims that hormone levels in parents are influential for determining their children's sex. NB female's digit ratio, but not males', was associated to the level of AF testosterone. The mother's 2D:4D ratios were positively correlated with their daughters' 2D:4D, but the same was not observed for male NBs, suggesting that prenatal testosterone levels in male fetus lead their 2D:4D ratios to stray from their mothers' with high individual variability.

Candida albicans PROTEIN PROFILE CHANGES IN RESPONSE TO THE BUTANOLIC EXTRACT OF Sapindus saponariaL.

Candida albicans is an opportunistic human pathogen that is capable of causing superficial and systemic infections in immunocompromised patients. Extracts of Sapindus saponaria have been used as antimicrobial agents against various organisms. In the present study, we used a combination of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify the changes in protein abundance of C. albicans after exposure to the minimal inhibitory concentration (MIC) and sub-minimal inhibitory concentration (sub-MIC) of the butanolic extract (BUTE) of S. saponaria and also to fluconazole. A total of six different proteins with greater than 1.5 fold induction or repression relative to the untreated control cells were identified among the three treatments. In general, proteins/enzymes involved with the glycolysis (GPM1, ENO1, FBA1), amino acid metabolism (ILV5, PDC11) and protein synthesis (ASC1) pathways were detected. In conclusion, our findings reveal antifungal-induced changes in protein abundance of C. albicans. By using the previously identified components of the BUTE of S. saponaria(e.g., saponins and sesquiterpene oligoglycosides), it will be possible to compare the behavior of compounds with unknown mechanisms of action, and this knowledge will help to focus the subsequent biochemical work aimed at defining the effects of these compounds.

Characterization of sol-gel matrices with entrapped cutinase

Dissertação para obtenção do Grau de Doutor em Engenharia Química e Bioquímica

Cortisol e testosterona no plasma da grávida e no líquido amniótico: variação com o relaxamento e correlação com o rácio digital 2D:4D no recém-nascido

RESUMO: Conclusão - Este estudo sugere que mulheres grávidas beneficiam da prática rotineira de 30 min de relaxamento perante a eminência de um acontecimento indutor de stress. O benefício será tanto psicológico como fisiológico e será mais marcado de manhã do que de tarde. As mulheres com maior tendência ansiosa, tipicamente as mais jovens e com menos idade gestacional, serão as que mais beneficiam. Entre os tipos de intervenção experimentados, todos eles não dispendiosos, a audição de música foi globalmente o mais eficaz para o propósito pretendido. A variação do cortisol e a variação da testosterona estão altamente correlacionadas, e este efeito poderá ser explorado em futuras intervenções relaxantes. Programas curtos de relaxamento poderão conduzir a benefícios para o bem-estar psicológico e para a saúde da grávida, com benefícios adicionais no desfecho da gravidez. O rácio digital 2D:4D do recém-nascido apresenta características sexualmente dimórficas, sendo mais baixo em rapazes que em raparigas, confirmando a influência dos androgénios pré-natais, mas há grande sobreposição entre os sexos pelo que, isoladamente, não pode ser utilizado como marcador de risco. No entanto, se associado a outros marcadores biológicos, como o peso ao nascer e a circunferência cerebral, entre outros, poderá ser um parâmetro a valorizar desde cedo, possivelmente no acompanhamento pediátrico. Nas raparigas, o RD da mãe e a testosterona no LA explicam uma proporção significativa do seu 2D:4D. Em ambos os sexos, o rácio 2D:4D aumenta após o nascimento e o dimorfismo sexual observado com base em amostras de crianças com mais de dois anos de idade, é afectado pela testosterona pós-natal, especialmente no que se refere à mão direita.----------ABSTRACT: Conclusions - This study suggests that pregnant women would benefit from the routine practice of 30min relaxation when at the imminence of a stressful event. The benefit would be both psychological and physiological and would be more pronounced in the morning than in the afternoon. Women tending to be more anxious, typically younger and at a lower gestational age, are the ones that benefit the most. Among the interventions investigated, all being non-expensive and easy to implement, listening to relaxing music was globally the most efficient for the benefit intended. The variation of cortisol and the variation of testosterone are significantly correlated, an association that might be used in future relaxation interventions. It is feasible to conceive short relaxation programs aimed at benefiting psychological well-being and health in pregnancy, with additional benefits for pregnancy outcome. The digital ratio 2D:4D is sexually dimorphic, being lower in males than in females, confirming the influence of pre-natal androgens. There is however large overlap between sexes, rendering the 2D:4D a risk marker not to be used in isolation but rather in association with other markers, like for instance the weight at birth and the head circumference, wherein it may be a relevant parameter for pediatric follow-up. In newborn females, a significant amount of variability in 2D:4D is accounted for by their mother’s digital ratio and AF testosterone. In both sexes, the 2D:4D ratio increases following birth and sexual dimorphism,as assessed from samples of subjects with more than 2 years of age, is affected by post-natal testosterone, especially in what concerns the right hand.

Monitoring and modelling of membrane bioreactors for wastewater treatment incorporating 2D fluorescence spectroscopy

Dissertação para obtenção do Grau de Doutor em Engenharia Química e Bioquímica, Especialidade em Engenharia Bioquímica

2D position system for a mobile robot in unstructured environments

Nowadays, several sensors and mechanisms are available to estimate a mobile robot trajectory and location with respect to its surroundings. Usually absolute positioning mechanisms are the most accurate, but they also are the most expensive ones, and require pre installed equipment in the environment. Therefore, a system capable of measuring its motion and location within the environment (relative positioning) has been a research goal since the beginning of autonomous vehicles. With the increasing of the computational performance, computer vision has become faster and, therefore, became possible to incorporate it in a mobile robot. In visual odometry feature based approaches, the model estimation requires absence of feature association outliers for an accurate motion. Outliers rejection is a delicate process considering there is always a trade-off between speed and reliability of the system. This dissertation proposes an indoor 2D position system using Visual Odometry. The mobile robot has a camera pointed to the ceiling, for image analysis. As requirements, the ceiling and the oor (where the robot moves) must be planes. In the literature, RANSAC is a widely used method for outlier rejection. However, it might be slow in critical circumstances. Therefore, it is proposed a new algorithm that accelerates RANSAC, maintaining its reliability. The algorithm, called FMBF, consists on comparing image texture patterns between pictures, preserving the most similar ones. There are several types of comparisons, with different computational cost and reliability. FMBF manages those comparisons in order to optimize the trade-off between speed and reliability.

Paracoccidioidomycosis: no genetic damage in human peripheral blood cells of patients assessed by single-cell gel (comet) assay

Paracoccidioidomycosis is a systemic fungal infection caused by Paracoccidioides brasiliensis. As infectious diseases can cause DNA damage, the authors aimed at analyzing DNA breakage in peripheral blood cells of patients with paracoccidioidomycosis by using the comet assay. The results suggested that paracoccidioidomycosis does not cause genotoxicity.

Modelação automática 2D de mineralizações filonianas – aplicação às Minas da Panasqueira

Neste trabalho apresenta-se uma metodologia protótipo destinada a efectuar a modelação automática a 2D da morfologia de mineralizações filonianas. Em primeiro lugar procede-se à estimação do número de ocorrências de filões em cada bloco, de uma malha de blocos, em que se subdividiu o volume em estudo. Esta intensidade de ocorrências é quantificada por uma variável que representa o número de filões por metro linear(NFM) intersectados por uma sondagem vertical, e constitui o objectivo de intensidade a atingir. Seguidamente são gerados por simulação, linhas poligonais ou arcos correspondentes ao posicionamento dos filões no perfil. Estes arcos ligam alguns pares de intersecções de filões amostrados nas sondagens e são escolhidos ao acaso segundo regras de orientação e distância. Seguidamente faz-se a avaliação da intensidade local de filões do modelo e, para os locais onde exista défice de filões relativamente ao objectivo, são adicionadas intersecções virtuais, ou seja, que não foram reconhecidas por sondagens. Este procedimento continua até o modelo dos filões se aproximar do objectivo previamente definido. O conjunto dos arcos em cada perfil, associado às espessuras de intersecção observadas nas sondagens, constitui um modelo morfológico dos filões em estrutura vectorial. No final faz-se a avaliação quantitativa do modelo e da incerteza. Os dados do estudo prático que motivaram o desenvolvimento da metodologia foram recolhidos no depósito mineral das Minas da Panasqueira. Os resultados obtidos no estudo mostram que a introdução de uma metodologia automática para a modelação vectorial de filões mineralizados é uma mais-valia, porque permite gerar modelos mais realistas e com melhor controlo geológico e resolução do que as abordagens clássicas de possança e acumulação, constituindo um auxiliar precioso de avaliação de reservas de minério.

Proteómica microbiana: análise comparativa da bactéria redutora de sulfato Desulfovibrio desulfuricans ATCC 27774 e das actinobactérias Salinispora arenicola e Salinispora pacifica

As bactérias redutoras de sulfato (BRS) são um grupo diversificado de micro-organismos anaeróbios que obtêm energia a partir da redução dissimilativa do sulfato. Algumas espécies de BRS possuem versatilidade a nível respiratório, como é o caso de Desulfovibrio desulfuricans ATCC 27774, devido à utilização de aceitadores finais de eletrões alternativos. Neste contexto, o objetivo da primeira parte desta dissertação consistiu na identificação das ferramentas metabólicas (proteínas) envolvidas na flexibilidade respiratória desta bacteria induzidas por diferentes aceitadores de eletrões (nitrato vs sulfato). Assim, os extratos proteicos totais de células de D. desulfuricans crescidas em meios VMN com nitrato ou sulfato, foram analisados por eletroforese bidimensional (2DE), num gradiente de pH 4 - 7. Nas condições experimentais testadas, foram observados 604 e 519 spots de proteínas nos géis de células crescidas em meio contendo nitrato ou sulfato, respetivamente. Pela avaliação estatística foi possível observar aproximadamente 25 % de spots diferenciais. Os resultados obtidos sugerem que na presença de nitrato, a bactéria não só cresce mais rapidamente e com maior rendimento, como também produz uma maior quantidade de proteínas. Estes dados foram relacionados com a adaptação de D. desulfuricans ao substrato respiratório alternativo. Tal como esperado, nos ensaios das atividades enzimáticas das redutases do nitrito e do nitrato, foi possível observar maior atividade nos extratos das células crescidas em nitrato do que em sulfato. As actinobactérias marinhas do género Salinispora, têm vindo a ser exploradas como fontes de biofármacos naturais. Assim, na segunda parte deste trabalho, realizou-se um estudo preliminar, que pretendeu caracterizar as proteínas envolvidas na biossíntese destes compostos bioactivos em S. arenicola e de S. pacifica. Para tal, foi utilizada uma abordagem proteómica diferencial, baseada em 2DE. Surpreendentemente, os perfis proteicos das duas espécies mostraram-se bastante distintos, tendo-se identificado apenas 37 spots comuns entre os 650 observados no gel de S. arenicola e 510 no gel de S. pacifica.

Mechanical characterization of aortas using 2D ultrasound elastography

Rupture of aortic aneurysms (AA) is a major cause of death in the Western world. Currently, clinical decision upon surgical intervention is based on the diameter of the aneurysm. However, this method is not fully adequate. Noninvasive assessment of the elastic properties of the arterial wall can be a better predictor for AA growth and rupture risk. The purpose of this study is to estimate mechanical properties of the aortic wall using in vitro inflation testing and 2D ultrasound (US) elastography, and investigate the performance of the proposed methodology for physiological conditions. Two different inflation experiments were performed on twelve porcine aortas: 1) a static experiment for a large pressure range (0 – 140 mmHg); 2) a dynamic experiment closely mimicking the in vivo hemodynamics at physiological pressures (70 – 130 mmHg). 2D raw radiofrequency (RF) US datasets were acquired for one longitudinal and two cross-sectional imaging planes, for both experiments. The RF-data were manually segmented and a 2D vessel wall displacement tracking algorithm was applied to obtain the aortic diameter–time behavior. The shear modulus G was estimated assuming a Neo-Hookean material model. In addition, an incremental study based on the static data was performed to: 1) investigate the changes in G for increasing mean arterial pressure (MAP), for a certain pressure difference (30, 40, 50 and 60 mmHg); 2) compare the results with those from the dynamic experiment, for the same pressure range. The resulting shear modulus G was 94 ± 16 kPa for the static experiment, which is in agreement with literature. A linear dependency on MAP was found for G, yet the effect of the pressure difference was negligible. The dynamic data revealed a G of 250 ± 20 kPa. For the same pressure range, the incremental shear modulus (Ginc) was 240 ± 39 kPa, which is in agreement with the former. In general, for all experiments, no significant differences in the values of G were found between different image planes. This study shows that 2D US elastography of aortas during inflation testing is feasible under controlled and physiological circumstances. In future studies, the in vivo, dynamic experiment should be repeated for a range of MAPs and pathological vessels should be examined. Furthermore, the use of more complex material models needs to be considered to describe the non-linear behavior of the vascular tissue.

Caracterização de cepas de referência de Leptospira sp utilizando a técnica de pulsed field gel electrophoresis

INTRODUÇÃO: A leptospirose é uma zoonose endêmica, mundialmente distribuída, causada por bactérias do gênero Leptospira. Este gênero compreende espécies patogênicas e saprofíticas, com mais de 200 sorovares distintos, dificultando sua caracterização. A técnica de pulsed field gel electrophoresis tem sido empregada como uma ferramenta para auxiliar nesta caracterização. Os objetivos deste trabalho foram padronizar a técnica de PFGE, determinar os perfis moleculares das cepas de referência utilizadas pelo Laboratório de Referência Nacional para Leptospirose/Centro Colaborador da Organização Mundial de Saúde para Leptospirose e criar um banco de dados com estes perfis. MÉTODOS: Foram analisadas, por PFGE, dezenove cepas utilizando a enzima de restrição NotI. RESULTADOS: Cada cepa apresentou um perfil único que pode ser considerado como uma identidade genômica específica, com exceção dos sorovares Icterohaemorrhagiae e Copenhageni, cujos perfis foram indistinguíveis. CONCLUSÕES: Dessa forma, foi possível a criação de um banco de perfis moleculares que está sendo utilizado no Laboratório para a comparação e identificação de cepas isoladas de quadros clínicos.

Sol-gel entrapped biosystems: enzyme(s) and whole cells

In this work two different procedures to utilize the sol-gel technology were applied to immobilize/encapsulate enzymes and living cells. CO2 has reached levels in the atmosphere that make it a pollutant. New methods to utilize this gas to obtain products of added value can be very important, both from an environmentally point of view and from an economic standpoint. The first goal of this work was to study the first reaction of a sequential, three-step, enzymatic process that carries out the conversion of CO2 to methanol. Of the three oxidoreductases involved, our focus was on formate dehydrogenase (FateDH) that converts CO2 to formate. This reaction requires the presence of the cofactor β-nicotinamide adenine dinucleotide in reduced form (NADH). The cofactor is expensive and unstable. Our experiments were directed towards generating NADH from its oxidized form (NAD+), using glutamate dehydrogenase (GDH). The formation of NADH from NAD+ in aqueous medium was studied with both free and sol-gel entrapped GDH. This reaction was then followed by the conversion of CO2 to formate, catalysed by free or sol-gel entrapped FateDH. The quantification of NADH/NAD+ was made using UV/Vis spectroscopy. Our results showed that it was possible to couple the GDH-catalyzed generation of the cofactor NADH with the FateDH-catalyzed conversion of CO2, as confirmed by the detection of formate in the medium, using High Performance Liquid Chromatography (HPLC). The immobilization of living cells can be advantageous from the standpoint of ease of recovery, reutilization and physical separation from the medium. Also dead cells may not always exhibit enzymatic activities found with living cells. In this work cell encapsulation was performed using Escherichia coli bacteria. To reduce toxicity for living organisms, the sol-gel method was different than for enzymes, and involved the use of aqueous-based precursors. Initial encapsulation experiments and viability tests were carried out with E. coli K12. Our results showed that sol-gel entrapment of the cells was achieved, and that cell viability could be increased with additives, namely betaine that led to greater viability improvement and was selected for further studies. For an approach to “in-cell” Nuclear Magnetic Resonance (NMR) experiments, the expression of the protein ctCBM11 was performed in E. coli BL21. It was possible to obtain an NMR signal from the entrapped cells, a considerable proportion of which remained alive after the NMR experiments. However, it was not possible to obtain a distinctive NMR signal from the target protein to distinguish it from the other proteins in the cell.