Backside-surface imprinting as a new strategy to generate specific plastic antibody materials

A backside protein-surface imprinting process is presented herein as a novel way to generate specific synthetic antibody materials. The template is covalently bonded to a carboxylated-PVC supporting film previously cast on gold, let to interact with charged monomers and surrounded next by another thick polymer. This polymer is then covalently attached to a transducing element and the backside of this structure (supporting film plus template) is removed as a regular “tape”. The new sensing layer is exposed after the full template removal, showing a high density of re-binding positions, as evidenced by SEM. To ensure that the templates have been efficiently removed, this re-binding layer was cleaned further with a proteolytic enzyme and solution washout. The final material was named MAPS, as in the back-side reading of SPAM, because it acts as a back-side imprinting of this recent approach. It was able to generate, for the first time, a specific response to a complex biomolecule from a synthetic material. Non-imprinted materials (NIMs) were also produced as blank and were used as a control of the imprinting process. All chemical modifications were followed by electrochemical techniques. This was done on a supporting film and transducing element of both MAPS and NIM. Only the MAPS-based device responded to oxLDL and the sensing layer was insensitive to other serum proteins, such as myoglobin and haemoglobin. Linear behaviour between log(C, μg mL−1) versus charged tranfer resistance (RCT, Ω) was observed by electrochemical impedance spectroscopy (EIS). Calibrations made in Fetal Calf Serum (FCS) were linear from 2.5 to 12.5 μg mL−1 (RCT = 946.12 × log C + 1590.7) with an R-squared of 0.9966. Overall, these were promising results towards the design of materials acting close to the natural antibodies and applied to practical use of clinical interest

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

Employing bacteria machinery for antibiotic detection: using DNA gyrase for ciprofloxacin detection

This work describes a new successful approach for designing biosensors that detect antibiotics. It makes use of a biomimetic strategy, by employing the biochemical target of a given antibiotic as its biorecognition element. This principle was tested herein for quinolones, which target DNA gyrase in bacteria. Ciprofloxacin (CIPRO) was tested as a representative antibiotic from the quinolone group; the sensitivity of biosensor to this group was confirmed by checking the response to another quinolone antibiotic (norfloxacin, NOR) and to a non-quinolone antibiotic (ampicillin, AMP). The biorecognition element used was DNA gyrase attached by ionic interactions to a carbon support, on a working electrode on common screen-printed electrodes (SPEs). The response against antibiotics was tested for increasing concentrations of CIPRO, NOR or AMP, and following the subsequent electrical changes by electrochemical impedance spectroscopy. The DNAgyrase biosensor showed sensitive responses for CIPRO and NOR, for concentrations down to 3.02nM and 30.2nM, respectively, with a very wide response range for CRIPRO, up to 30.2µM. Its response was also confirmed selective for quinolones, when compared to its response against AMP. Further comparison to an immunosensor of similar design (adding antibodies instead of DNA gyrase) was made, revealing favourable features for the new biomimetic biosensor with 1.52nM of limit of detection (LOD). Overall, the new approach presented herein is simple and effective for antibiotic detection, displaying a selective response against a given antibiotic group. The use of bacterial machinery as biorecognition element in biosensors may also provide a valuable tool to study the mechanism of action in bacterial cells of new drugs. This is especially important in the development of new drugs to fight bacterial resistance.The authors acknowledge funding from project PTDC/AAG-TEC/5400/2014 funded by European funds, through FEDER (European Funding or Regional Development) via COMPETE2020 – POCI (operational program for internationalization and competitively) and by national funding through the National Foundation for Science and Technology, I.P. (FCT). ARC also acknowledge funding to National Foundation for Science and Technology, I.P., through the PhD Grant, SFRH/BD/130107/2017.info:eu-repo/semantics/publishedVersio

Vaccination Against Amyloidogenic Aggregates in Pancreatic Islets Prevents Development of Type 2 Diabetes Mellitus

Type 2 diabetes mellitus (T2DM) is a chronic progressive disease characterized by insulin resistance and insufficient insulin secretion to maintain normoglycemia. The majority of T2DM patients bear amyloid deposits mainly composed of islet amyloid polypeptide (IAPP) in their pancreatic islets. These-originally β-cell secretory products-extracellular aggregates are cytotoxic for insulin-producing β-cells and are associated with β-cell loss and inflammation in T2DM advanced stages. Due to the absence of T2DM preventive medicaments and the presence of only symptomatic drugs acting towards increasing hormone secretion and action, we aimed at establishing a novel disease-modifying therapy targeting the cytotoxic IAPP deposits in order to prevent the development of T2DM. We generated a vaccine based on virus-like particles (VLPs), devoid of genomic material, coupled to IAPP peptides inducing specific antibodies against aggregated, but not monomeric IAPP. Using a mouse model of islet amyloidosis, we demonstrate in vivo that our vaccine induced a potent antibody response against aggregated, but not soluble IAPP, strikingly preventing IAPP depositions, delaying onset of hyperglycemia and the induction of the associated pro-inflammatory cytokine Interleukin 1β (IL-1β). We offer the first cost-effective and safe disease-modifying approach targeting islet dysfunction in T2DM, preventing pathogenic aggregates without disturbing physiological IAPP function.These studies were funded by a project grant from the Swiss National Foundation (SNF). We acknowledge the technical assistance of Sydney W. Pence and Faith Slubowski at the Institute of Veterinary Physiology, University of Zürich. We appreciate the kind possibility given by Nanolive (Lausanne, Switzerland) for the opportunity and the collaborative acquisition of tomographic pictures.S

The use of a new calcium mesoporous silica nanoparticle versus calcium and/or fluoride products in reducing the progression of dental erosion

Objective: There is increasingly common the consumption more times a day of foods and acidic drinks in the diet of the population. The present study aimed to evaluate and compare the effects of a calcium mesoporous silica nanoparticle single application of other calcium and/or fluoride products in reducing the progression of dental erosion. Methodology: Half of the eroded area was covered of 60 blocks of enamel, after which the block was submitted to the following treatments: (Ca2+-MSN), casein phosphopeptide–amorphous calcium phosphate (CPP-ACP); CPP-ACP/F-(900 ppm F−); titanium tetrafluoride (TiF4 1%) (positive control); sodium fluoride (NaF 1.36%) (positive control); and Milli-Q® water (negative control) before being submitted to a second erosive challenge. A surface analysis was performed via a three-dimensional (3D) noncontact optical profilometry to assess the volumetric roughness (Sa) and tooth structure loss (TSL) and and through scanning electron microscopy (MEV). An analysis of variance (ANOVA) and Tukey’s test were performed. Results: Regarding Sa, all experimental groups exhibited less roughness than the control (p<0.05). The TSL analysis revealed that the Ca2+-MSN and NaF groups were similar (p>0.05) and more effective in minimizing tooth loss compared with the other groups (p<0.05). Conclusions: The Ca2+-MSN and NaF treatments were superior compared with the others and the negative control

Preclinical development of a vaccine against oligomeric alpha-synuclein based on virus-like particles

Parkinson's disease (PD) is a progressive and currently incurable neurological disorder characterised by the loss of midbrain dopaminergic neurons and the accumulation of aggregated alpha-synuclein (a-syn). Oligomeric a-syn is proposed to play a central role in spreading protein aggregation in the brain with associated cellular toxicity contributing to a progressive neurological decline. For this reason, a-syn oligomers have attracted interest as therapeutic targets for neurodegenerative conditions such as PD and other alpha-synucleinopathies. In addition to strategies using small molecules, neutralisation of the toxic oligomers by antibodies represents an attractive and highly specific strategy for reducing disease progression. Emerging active immunisation approaches using vaccines are already being trialled to induce such antibodies. Here we propose a novel vaccine based on the RNA bacteriophage (Qbeta) virus-like particle conjugated with short peptides of human a-syn. High titres of antibodies were successfully and safely generated in wild-type and human a-syn over-expressing (SNCA-OVX) transgenic mice following vaccination. Antibodies from vaccine candidates targeting the C-terminal regions of a-syn were able to recognise Lewy bodies, the hallmark aggregates in human PD brains. Furthermore, antibodies specifically targeted oligomeric and aggregated a-syn as they exhibited 100 times greater affinity for oligomeric species over monomer a-syn proteins in solution. In the SNCA-OVX transgenic mice used, vaccination was, however, unable to confer significant changes to oligomeric a-syn bioburden. Similarly, there was no discernible effect of vaccine treatment on behavioural phenotype as compared to control groups. Thus, antibodies specific for oligomeric a-syn induced by vaccination were unable to treat symptoms of PD in this particular mouse model.</p

Avaliação de adaptação e molhamento de resinas de diferentes viscosidades à parede de fundo de cavidades Classe I / Wetting and wetting capacity of composite resins with different viscosities to Class I internal walls

Este estudo avaliou a capacidade de molhamento e adaptação compósitos resinosos de três diferentes viscosidades às paredes cavitárias de preparos padronizados realizados em dentes bovinos simulando cavidades tipo Classe I de Black. Um molde realizado com silicone de adição foi feito de cada cavidade previamente à restauração para título de controle da capacidade de molhamento e adaptação. Com as restaurações finalizadas, os dentes foram submetidos à desmineralização de sua estrutura permitindo que a restauração fosse facilmente destacada para a sua avaliação. Os corpos de prova foram avaliados com o auxílio de um perfilômetro 3D. Dentre os grupos estudados o melhor desempenho foi da resina bulk fill flow. Os resultados sugerem que a viscosidade dos compósitos resinosos restauradores interfere diretamente na capacidade de molhamento e adaptação deles às paredes cavitarias e que devem ser considerados pelo menos como uma camada de aplicação inicial

Correction: Zika virus-derived e-diii protein displayed on immunologically optimized vlps induces neutralizing antibodies without causing enhancement of dengue virus infection. vaccines 2019, 7, 72

The authors wish to make the following correction to their paper [1]. The same image was mistakenly selected for Figures 2 and 3. The image became replaced as you see in Figure 3 below

Tipos de plantio e fertilizante biológico no cafeeiro em função do índice térmico

The implantation phase of coffee is one of the most important stages of the crop. It is worth mentioning that the different implantation systems provide different conditions for the development of crops. In addition, in order to achieve a good initial development of the coffee, the producers have applied in newly transplanted crops organic fertilizers, which has the advantage of improving soil biota, as well as reducing costs with the acquisition of mineral fertilizers. Few researches have jointly tested the efficiency of the different systems of implantation and the biological fertilization in the coffee tree. The objective of this study was to test the efficacy of different types of planting and of the biological fertilizer on the initial growth of arabica coffee, as a function of the thermal index. The experiment was developed in southern Minas Gerais, Brazil. The experimental design was in randomized blocks, in a 3 x 2 factorial scheme with 4 replicates, in a total of 6 treatments. The following were used: a) three systems of implantation (conventional planting, no-tillage in the pit and minimum cultivation); and b) two doses of the biological fertilizer (presence and absence). The growth parameters and the first coffee crop were evaluated. The evaluations were carried out according to the degree days counted from the date of planting. The results were submitted to analysis of variance. The means were grouped by the Scott-Knott method. The coffee plants implanted in the planting system “in pits” showed the best results, both in vegetative parameters and in productivity. The use of the biological fertilization in the newly transplanted arabica coffee tree did not promote improvements in the vegetative growth and first coffee production.A fase de implantação do cafeeiro consiste em uma das etapas mais importantes da cultura. Vale ressaltar, que os diferentes sistemas de implantação proporcionam condições distintas de desenvolvimento das culturas. Além disso, visando um bom desenvolvimento inicial do cafeeiro, os produtores têm aplicado em lavouras recém-transplantadas adubos biológicos, que têm como vantagem a melhora da biota do solo, além da redução de custos com a aquisição de fertilizantes minerais. Poucas pesquisas têm testado de forma conjunta a eficiência dos diferentes sistemas de implantação e a adubação biológica no cafeeiro. Assim, objetivou-se testar a eficácia de diferentes tipos de plantio e do fertilizante biológico no crescimento inicial do cafeeiro arábica, cultivar Catucaí, em função do índice térmico. O experimento foi desenvolvido no Sul de Minas Gerais, Brasil. O delineamento experimental foi em blocos casualizados, em esquema fatorial 3 x 2 com 4 repetições, num total de 6 tratamentos. Foram utilizados: a) três sistemas de implantação (plantio convencional, o plantio direto na cova e cultivo mínimo); e b) duas doses do adubo biológico (presença e ausência). Foram avaliados os parâmetros de crescimento e a primeira safra do cafeeiro. As avaliações foram realizadas em função dos graus dias contabilizados a partir da data de plantio. Os resultados foram submetidos à análise de variância e as médias agrupadas pelo teste de Scott-Knott. As plantas de café implantadas no sistema de plantio “em covas” evidenciaram os melhores resultados, tanto nos parâmetros vegetativos como na produtividade. A utilização da adubação biológica no cafeeiro arábica recém-transplantado não promoveu melhorias no crescimento vegetativo e primeira produção do cafeeiro

Estudo do comportamento mecânico da dureza e da morfologia do compósito Al2O3-YAG

Os óxidos cerâmicos possuem alta resistência à oxidação e à corrosão em ambientes agressivos e em elevadas temperaturas, o que torna o seu emprego bastante atraente em relação a outros cerâmicos. Na década de 90, diversos pesquisadores mostraram o YAG como sendo o óxido de maior resistência à fluência em elevadas temperaturas. Nesta pesquisa, os pós reagentes, Al2O3 e Y2O3 foram misturados nas proporções em peso de 63,65 e 36,35% e 80,00 e 20,00%, respectivamente. Após, a mistura foi moída em moinho de bolas, por 5h, secos em estufa por 48 h, à 120 °C. Depois desaglomerados em gral e pistilo e peneiramento em peneiras de 120 mesh. Na etapa de compactação, os pós foram prensados em matriz e punção de aço, à 70 MPa, por 20 s de aplicação de carga. No processo de sinterização, foram realizados em forno em 1500 e 1600 °C, com 3 h de patamar, com taxa de aquecimento e resfriamento de 5 ºC/min, ao ar. Foram produzidos cinco corpos de prova para cada condição de sinterização. As amostras sinterizadas foram cortadas a fim de serem embutidas em baquelite. Após esta etapa as amostras foram lixadas em lixas diamantadas e foram polidas em pasta de diamante. As amostras foram avaliadas por dureza por microindentação Vickers e por Microscopia Eletrônica de Varredura (MEV). A amostra na composição eutética sinterizada à 1600°C apresentou maior dureza de 1200 HV.