Mechanical Properties of Mini-Implants Used in Extra-Radicular Anchorage

ABSTRACT Objective: To evaluate the mechanical properties of mini-implants (MIs) manufactured from stainless steel and compare them with conventional titanium-aluminum-vanadium alloy MIs. Material and Methods: The following groups were formed: G1 (n=24), 8×1.5 mm steel MIs; G2 (n=24), 12×2.0 mm steel MIs; and G3 (n=24), 10×1.5 mm titanium MIs. The 72 MIs were inserted in the infra zygomatic crest region of the maxilla and retromolar trigone in the jaw of 10 pigs. Pull-out, insertion torque, fracture and percussion tests were performed in order to measure the tensile strength, primary stability and fracture strength of MIs. A digital torque gauge was used to measure insertion and fracture torque, a universal mechanical testing machine was used for pull-out testing and a periotest device was used to measure the micromovement of MIs. For morphological and MI component evaluation, scanning electron microscopy (SEM) was performed. D'Agostino & Pearson, Kruskal-Wallis, and Dunn post-hoc and normality tests were used. Results: G2 insertion and fracture torques were significantly higher than G1 and G3 insertion and fracture torques (p<0.05). The pull-out and percussion tests presented similar values among the groups. SEM revealed that the fracture point was predominantly on the fourth thread for steel MIs (G1 and G2) and on the seventh thread for titanium-aluminum-vanadium MIs (G3). Conclusion: The mechanical properties of stainless steel MIs are superior to those of titanium-aluminum-vanadium alloy MIs.

Effect of Glycolic Acid, Maleic Acid, and EDTA in the Removal of Smear Layer from Root Canal Dentin

ABSTRACT Objective: To assess the efficacy of 5% Glycolic Acid (GA), 17% ethylenediaminetetraacetic acid (EDTA) and 7% maleic acid (MA), in removing the smear layer (SL). Material and Methods: For the experiment, forty single-rooted human teeth were selected. To perform the chemo‑mechanical preparation, the root canals were instrumented to an apical size of #30, along with simultaneous irrigation of 2.5% NaOCl. The samples were allotted to the experimental groups based on the final irrigating solution (n=10): (1) The GA group: 05%, (2) the EDTA group: 17%, (3) the MA group: 7%, and (4) the control group: Distilled water. The teeth were evaluated for the presence or absence of SL using SEM. Results: Comparing the 5% GA, 7% MA and 17% EDTA groups, no statistically significant differences were found at the coronal and middle thirds (p>0.05). However, in the apical third, MA had greater efficacy than EDTA (p=0.002) and GA (p=0.041), with a significant difference in the SL removal, while there was no significant difference between the latter (p=0.148). Conclusion: Thus, it was concluded that 7% maleic acid as a final irrigating solution is more efficacious than 17% EDTA and 5% glycolic acid in eliminating the smear layer from the apical portion of the root canal.

Avaliação das características de superfície e variação de massa de diferentes tipos de resina após escovação simulada / Evaluation of surface characteristics and mass variation of different types of resin after simulated toothbrushing

Introdução: O aumento da exigência estética pelos pacientes e o contínuo desenvolvimento de materiais - tais como resinas bulk fill de alta e baixa viscosidade e flow, demanda uma crescente necessidade de estudos nesta área para avaliar as propriedades dos materiais quando submetidos ao desgaste ocasionado pela escovação dentária ao longo do tempo. Objetivo: Avaliar as características superficiais de restaurações antes e após o processo de escovação simulada e comparar os resultados entre os mesmos. Materiais e métodos: Foram confeccionados ao total 80 corpos de prova (7x4mm), sendo 20 para cada tipo de resina utilizada, na cor A1: Bulk Fill One (3M/ESPE, St. Paul, MN, USA), Bulk Fill Flow (3M/ESPE, St. Paul, MN, USA), Z350XT (3M/ESPE St. Paul, MN, USA) e Z350XT Flow (3M/ESPE, St. Paul, MN, USA). Após a confecção, os corpos de prova permaneceram por 24 horas imersos em água destilada em estufa a 37°C, momento no qual foi realizado o acabamento e polimento com a sequência de discos de lixa Praxis (TDV, Brasil). Metade dos corpos de prova foi submetida a 60,000 ciclos de escovação simulada, enquanto a outra metade foi o grupo controle (n=10). A superfície das restaurações foi avaliada por meio do Microscópio Eletrônico de Varredura (MEV), rugosidade superficial (Ra) e variação de massa (através de pesagem) das restaurações antes e após o processo de escovação simulada. Para análise estatística, foi utilizada a análise de variância ANOVA dois fatores e pós teste de Tukey. Resultados: Na análise feita pelo MEV, em amplificação de 500x, foi observada a protuberância de partículas médias e pequenas em todas as resinas após a escovação simulada, com exceção da resina de tipo Z350 flow. Todos os materiais apresentaram aumento da Ra após a escovação simulada. Com relação a variação de massa, não houve diferença estatisticamente significativa. Conclusão: O processo abrasivo ocasionado pela escovação leva a mudanças na disposição das partículas, o que ocasiona mudanças na superfície do material e aumento na rugosidade. Por outro lado, não foi demonstrada diferença com relação a variação de massa (AU).

Introduction: The increasing of the aesthetic exigency by the patients and the continual development of materials ­ such as bulk fill and flow composites, demands a crescent need of studies on this area to evaluate the properties of the materials when submitted to the wear occasioned by toothbrushing abrasion over time. Objective: To evaluate the surface characteristics of restorations performed before and after the process of simulated toothbrushing and to compare the results between the composites. Materials and methods: In total, 80 samples were made (7x4mm), being 20 to each type of resin utilized, color A1: Bulk Fill One (3M/ESPE, St. Paul, MN, USA), Bulk Fill Flow (3M/ESPE, St. Paul, MN, USA), Z350XT (3M/ESPE St. Paul, MN, USA) and Z350XT Flow (3M/ESPE, St. Paul, MN, USA). After the confection of the specimens, the samples were immersed in distilled water for 24 hours and stored at 37°C in stove, moment that was performed the finishing and polishing with the sequence of sandpaper discs (TDV, Brasil). Half of the specimens was submitted to 60,000 cycles of simulated toothbrushing, meanwhile the other half was the control group (n=10). The surface of the restorations was evaluated through Scanning Electronic Microscopy (SEM), surface roughness (Ra), and weight loss (through measurements), in two moments: before and after the process of simulated toothbrushing. To the statistical analysis, it was used the variance analysis ANOVA two factors and Tukey's post test. Results: In the analysis performed by SEM, in amplification of 500x, it was observed the presence of medium and small protruding particles in all resins tested after simulated toothbrushing, with exception of the resin Z350 flow. All materials presented increasing on Ra after simulated toothbrushing. Regarding to mass variation, there was not any significant statistical difference. Conclusion: The abrasive process occasioned by toothbrushing leads to changes on the particles disposition, which causes changes on the surface and increase on the roughness. On the other hand, there was not any difference regarding to mass variation (AU).

Efeitos do laser de baixa intensidade em osteoblastos cultivados em arcabouços nanofibrosos de ácido polilático / Effects of low-intensity laser on osteoblasts cultured on polylactic acid nanofibrous scaffolds

A engenharia de tecidos ósseos é um ramo importante da medicina regenerativa e envolve o desenvolvimento de arcabouços com composição e arquitetura favoráveis à integração celular, além do estudo de fatores capazes de promover a adesão e proliferação celular, incluindo estímulos químicos e biofísicos. O objetivo do estudo foi avaliar a utilização do laser de baixa intensidade (LBI) como uma ferramenta para promover a bioestimulação in vitro de células osteoblásticas cultivadas em arcabouços nanofibrosos de ácido polilático (PLA). Os arcabouços foram produzidos pela técnica de eletrofiação e caracterizados quanto à molhabilidade, composição pela espectroscopia no infravermelho por transformada de Fourier (FTIR), morfologia da superfície por microscópica eletrônica de varredura (MEV), caracterização termogravimétrica (TGA), calorimetria diferencial exploratória (DSC) e cristalinidade por difração de raios-X (DRX). Os ensaios biológicos foram conduzidos com osteoblastos da linhagem OFCOL II cultivados na superfície dos arcabouços e submetidos ou não (grupo controle) a irradiação com laser diodo InGaAIP na potência de 30 mW, nas doses de 1, 4 e 6 J/cm² e nos comprimentos de onda de 660 nm (grupos V1, V4, V6, respectivo as doses) e 780 nm (grupos I1, I4 e I6, respectivo as doses). Os efeitos do LBI na proliferação dos osteoblastos foram avaliados através do método bioquímico Alamar Blue, nos intervalos de 24, 48 e 72h, enquanto a viabilidade e a morfologia celular foram analisadas no intervalo de 72h, através do ensaio Live/Dead e da microscopia eletrônica de varredura (MEV), respectivamente. Os dados do ensaio bioquímico de Alamar Blue mostraram uma maior proliferação celular nos grupos V6 em todos os intervalos analíticos em comparação ao grupo controle (p<0,05). Outras diferenças entre o grupo controle e irradiados foram encontradas apenas nos intervalos de 48h e 72h para V1, e para o grupo IV6 em 72h. O ensaio Live/Dead revelou um aumento na viabilidade celular nos grupos trados com LBI, sendo significativamente maior no grupo V1 quando comparado ao grupo controle. A análise por MEV mostrou adequada interação dos osteoblastos aos arcabouços, com o corpo celular se espalhando ao longo do eixo da nanofibra e a presença de contatos físicos mais evidentes, através da formação de ligação por meio de filopódios e lamelipódios, nos grupos V1, V6 e I6. Em conjunto, os dados do presente trabalho mostraram que o LBI promove a bioestimulação de osteoblastos cultivados sobre nanofibras de PLA, o que aponta para o seu uso potencial nas técnicas de engenharia tecidual óssea, sobretudo no que se refere ao uso do comprimento de onda de 660 nm, a qual apresentou grupos com mais resultados significativos (AU).

Bone tissue engineering is a relevant branch of regenerative medicine and involves the development of scaffolds with composition and architecture favorable to cell integration, in addition to studying factors capable of promoting cell adhesion and proliferation, including chemical and biophysical stimuli. The study aimed to evaluate the use of low-level laser irradiation (LLLI) to promote in vitro biostimulation of osteoblastic cells cultured on polylactic acid (PLA) nanofibrous scaffolds. The scaffolds were produced by the electrospinning technique and characterized in terms of wettability, composition by Fourier transform infrared spectroscopy (FTIR), surface morphology by scanning electron microscopy (SEM), thermogravimetric characterization (TGA), differential scanning calorimetry (DSC) and crystallinity by Xray diffraction (XRD). The biological assays were conducted with osteoblasts of the OFCOL II lineage cultured on the surface of the scaffolds and submitted or not (control group) to irradiation with InGaAIP diode laser, power of 30 mW, with doses of 1, 4 and 6 J/cm² and wavelengths of 660 nm (groups V1, V4, V6, respectively doses) and 780 nm (groups I1, I4 and I6, respectively doses). The effects of LLLT from the perspective of osteoblasts were evaluated using the biochemical method Alamar Blue assay, at intervals of 24, 48 and 72h, while cell viability and morphology were observed at 72h, using the Live/Dead assay and electron microscopy. scan (SEM), respectively. The Alamar Blue assay data showed more significant cell proliferation in groups in the V6 groups at all analytical intervals compared to the control group (p<0.05). Other differences between the control and irradiated groups were found only at intervals of 48h and 72h for V1, and for group IV6 at 72h. The Live/Dead assay revealed an increase in cell viability in the groups treated with LLLT, being significantly higher in the V1 group when compared to the control group. SEM analysis showed adequate interaction between osteoblasts and scaffolds, with the cell body spreading along the nanofiber axis and the presence of more evident physical contacts, through the formation of bonds through filopodia and lamellipodia, in groups V1, V6 and I6. Together, the data from the present study observed that LLLT promotes the biostimulation of osteoblasts cultured on PLA nanofibers, which pointed to its potential use in bone tissue engineering techniques, especially with regard to the use of the wavelength of 660 nm, which presented groups with more significant results (AU).

Research Techniques Made Simple: Volume Scanning Electron Microscopy.

Volume scanning electron microscopy (VSEM) involves the serial sectioning and imaging of a sample using scanning electron microscopy (SEM), followed by segmentation and three-dimensional (3D) reconstruction using computer software packages to allow visualization of 3D structures. VSEM can reveal qualitative and quantitative properties of organelles and cells within tissues at nanoscale. The ability to visualize spatial relationships of structures of interest within and across cells in 3D space in particular sets VSEM apart from conventional SEM and transmission electron microscopy. Here, we provide an overview of VSEM platforms and image processing, highlighting characteristics that will aid selection of a method to address specific research questions in dermatological research.

X-ray microscopy enables multiscale high-resolution 3D imaging of plant cells, tissues, and organs.

Capturing complete internal anatomies of plant organs and tissues within their relevant morphological context remains a key challenge in plant science. While plant growth and development are inherently multiscale, conventional light, fluorescence, and electron microscopy platforms are typically limited to imaging of plant microstructure from small flat samples that lack a direct spatial context to, and represent only a small portion of, the relevant plant macrostructures. We demonstrate technical advances with a lab-based X-ray microscope (XRM) that bridge the imaging gap by providing multiscale high-resolution three-dimensional (3D) volumes of intact plant samples from the cell to the whole plant level. Serial imaging of a single sample is shown to provide sub-micron 3D volumes co-registered with lower magnification scans for explicit contextual reference. High-quality 3D volume data from our enhanced methods facilitate sophisticated and effective computational segmentation. Advances in sample preparation make multimodal correlative imaging workflows possible, where a single resin-embedded plant sample is scanned via XRM to generate a 3D cell-level map, and then used to identify and zoom in on sub-cellular regions of interest for high-resolution scanning electron microscopy. In total, we present the methodologies for use of XRM in the multiscale and multimodal analysis of 3D plant features using numerous economically and scientifically important plant systems.

Obliteration of Dentinal Tubules by Desensitizing Agents Based on Silver Fluoride/Potassium Iodide or Pre-Reacted Glass Particles: An in Vitro Study

Abstract Objective: To evaluate the efficacy of desensitizing agents for the obliteration of dentinal tubules subjected or not to a simulated oral environment. Material and Methods: Dentinal discs (n=8) treated with Riva-Star (RS) or PRG-Barrier-Coat (PRG) were submitted (cycled) or not submitted (control) to erosive-abrasive-thermal cycles and evaluated using scanning electron microscopy/energy dispersive spectroscopic analysis. The variables analyzed were tubule obliteration and dentin surface chemical composition. Data were analyzed by non-parametric tests (p<0.05). Results: The cycled and control groups did not differ significantly for the responses in each material. The PRG control and cycled groups had fewer visible tubules and a higher proportion of totally obliterated tubules than the RS groups. The percentages of silver coverage were higher in the RS-control than in the RS-cycled. There was a significant inverse correlation between the presence of silver and non-obliterated tubules (R=-0.791; p<0.001). The percentages of carbon, aluminum, strontium, and potassium were significantly higher in the PRG-control and PRG-cycled compared to the RS control. The percentages of calcium, phosphorus, and silver were significantly higher in the RS compared to the PRG groups. PRG-control showed a higher percentage of boron than RS-control. Conclusion: PRG promoted greater tubule obliteration than SR. Simulated stress did not affect the obliterating effect of each agent. Greater silver coverage corresponded to a lower proportion of non-obliterated tubules in RS. Carbon, aluminum, strontium, boron, and potassium predominated in the dentin surface treated with PRG, while calcium, phosphorus, and silver prevailed in RS groups (AU).

Effect of Light-Curing Unit Type and Bulk-Fill Composite Resins with Different Photoinitiators on Marginal Gaps of Class II Restorations

Abstract Objective: To evaluate the effect of two types of light-curing units (second and third generations) and two types of bulk-fill composite resins with different photoinitiators - Tetric N-Ceram Bulk Fill (TNCB) and Xtra Fil (XTF) on gap formation at the gingival margins of Cl II restorations. Material and Methods: Fifty-six standard Cl II cavities were prepared on the mesial and distal surfaces of premolar teeth, with the gingival margin of the cavities 1 mm apical to the CEJ. The samples were randomly assigned to two groups based on the composite resin type and two subgroups based on the light-curing unit type and restored. After 5000 rounds of thermocycling, gingival margin gap in each sample was measured in µm under an electron microscope at ×2000 magnification. Data were analyzed by two-way ANOVA and Tukey tests (α=0.05). Results: Marginal gaps of TNCB composite resin were significantly smaller than those of XTF composite resin (p<0.001). There were no significant differences between the two light-curing units in each group (p=0.887 with XTF and p=0.999 with TNCB). Conclusion: The gaps at gingival margins of Cl II cavities with TNCB bulk-fill composite were smaller than XTF composite resin. Both composite resins can be cured with both the second- and third-generation LEDs (AU).

Caracterização mecânica, termodinâmica e química de fios CuNiTi ­ um estudo comparativo clínico e laboratorial / Mechanical, thermodynamic and chemical characterization of CuNiTi wires ­ a clinical and laboratory comparative study

Introdução: Os fios Copper Ni-Ti (CuNiTi) possuem indicações de uso clínico específicas, permanecendo no ambiente bucal por um longo período. Por esse motivo suas características mecânicas, termodinâmicas e estruturais devem ser preservadas durante todo o período de uso. Objetivos: Investigar se ocorrem alterações no comportamento mecânico, termodinâmico, estrutura e composição química superficial em fios CuNiTi 35°C submetidos ao envelhecimento in vivo (uso clínico) e in vitro (termociclagem), evidenciando as possíveis diferenças entre os métodos de envelhecimento. Material e Métodos: A amostra total foi constituída de 30 arcos pré-contornados 0.016", termodinâmicos com adição de cobre, e temperatura austenítica final (Af) de 35°C, da marca Ormco®. As análises destes fios resultaram em dois capítulos. Para o experimento clínico, 10 unidades de fios foram instaladas em 05 pacientes e permaneceram em meio bucal durante 30, 60 e 90 dias, e 05 fios foram analisados como recebidos (CR). A cada período, um hemiarco completo mais 1/3 (um terço) do hemiarco inferior direito foi retirado para análises. Para o experimento in vitro os 15 fios restantes foram submetidos à termociclagem para simulação do envelhecimento em ambiente oral durante 30 (600 ciclos), 60 (1200 ciclos) e 90 dias (1800 ciclos) com variação de temperatura entre 5°C e 55°C, com banhos de 90 segundos em cada temperatura, e transição de 15 segundos entre os banhos. Após cada período de envelhecimento in vivo e in vitro, os fios foram submetidos a um teste de padronização das suas dimensões através da medição dos diâmetros com paquímetro, Microscopia Eletrônica de Varredura (MEV), Espectrometria por Raios X Fluorescentes (FRX), ensaios de tração uniaxial e ensaio de Varredura Diferencial de Calorimetria (DSC). Resultados: No capítulo 1 a comparação entre os fios CR e os envelhecidos revelou que não houve diferença significativa entre os diâmetros das amostras, das forças obtidas ou mesmo das temperaturas Af, independentemente do tempo de permanência em boca. Foram encontrados predominantemente Ni, Ti, Cu e Al nas amostras, além de outros elementos químicos em concentrações variadas. No capítulo 2 não houve diferença significativa entre os diâmetros das amostras, das forças obtidas ou mesmo das temperaturas Af entre as amostras, independentemente do tempo ou do método de envelhecimento. Conclusão: As análises laboratoriais dos fios envelhecidos in vivo e in vitro foram comparadas, evidenciando que as características mecânicas, termodinâmicas e químicas dos fios permanecem estáveis mesmo até 90 dias de uso clínico ou 1800 ciclos de termociclagem. O método de envelhecimento in vitro se mostra uma alternativa viável para a análise destes parâmetros, em substituição aos métodos in vivo (AU).

Introduction: Copper Ni-Ti wires (CuNiTi) have specific indications for clinical use, remaining in the oral environment for a long period. For this reason, their mechanical, thermodynamic and structural characteristics must be preserved throughout the period of use of these materials. Objectives: To investigate whether there are changes in the mechanical, thermodynamic, structure and surface chemical composition of CuNiTi 35°C wires subjected to in vivo aging (clinical use) and in vitro (thermocycling), highlighting the possible differences between the aging methods. Material and Methods: The total sample was obtained from 30 pre-contoured Ormco® 0.016 thermodynamic archwires with copper addition and austenitic final temperature (Af) of 35°C. The analysis of these archwires resulted in two articles. For the clinical experiment, 10 units of wires were installed in 05 patients and remained in the oral environment for 30, 60 and 90 days, and 05 wires were analyzed as received. At each period, a complete hemiarch plus 1/3 of the right lower hemiarch was removed for analysis. For the in vitro experiment, the 15 remaining wires underwent thermocycling to simulate the aging of the oral environment for 30 (600 cycles), 60 (1200 cycles) and 90 days (1800 cycles) with a temperature range between 5°C and 55°C, with 90 second baths at each temperature, and 15 second transition between baths. After each in vivo and in vitro aging period, the wires underwent to a standardization test of their dimensions, by defining the diameters with a caliper, Scanning Electron Microscopy (SEM), Fluorescent X-Ray Spectrometry (FRX), tests of Uniaxial traction and Differential Scanning Calorimetry (DSC) test. Results: In chapter 1, the comparison between CR and aged wires revealed that there was no significant difference between the sample diameters, the forces obtained or even the temperatures Af, regardless the time spent in the mouth. Predominantly Ni, Ti, Cu and Al were found in the samples, in addition to other chemical elements in different concentrations. In chapter 2 there was no significant difference between the diameters of the samples, the forces obtained or even the temperatures Af between the samples, regardless of time or aging method. Conclusion: The comparison of the laboratoral analyzes of the archwires aged in vivo and in vitro showed that the mechanical, thermodynamic and the surface chemical characteristics were stable, even up to 90 days of clinical use or 1800 thermocycling cycles. The in vitro aging method is a viable alternative for the analysis of these parameters, replacing the in vivo methods (AU).

Exploring confocal laser scanning microscopy (CLSM) and fluorescence staining as a tool for imaging and quantifying traces of marine microbioerosion and their trace-making microendoliths.

Microscopic organisms that penetrate calcareous structures by actively dissolving the carbonate matrix, namely microendoliths, have an important influence on the breakdown of marine carbonates. The study of these microorganisms and the bioerosion traces they produce is crucial for understanding the impact of their bioeroding activity on the carbonate recycling in environments under global climate change. Traditionally, either the extracted microendoliths were studied by conventional microscopy or their traces were investigated using scanning electron microscopy (SEM) of epoxy resin casts. A visualisation of the microendoliths in situ, that is within their complex microbioerosion structures, was previously limited to the laborious and time-consuming double-inclusion cast-embedding technique. Here, we assess the applicability of various fluorescence staining methods in combination with confocal laser scanning microscopy (CLSM) for the study of fungal microendoliths in situ in partly translucent mollusc shells. Among the tested methods, specific staining with dyes against the DNA (nuclei) of the trace making organisms turned out to be a useful and reproducible approach. Bright and clearly delineated fluorescence signals of microendolithic nuclei allow, for instance, a differentiation between abandoned and still populated microborings. Furthermore, infiltrating the microborings with fluorescently stained resin seems to be of great capability for the visualisation and quantification of microbioerosion structures in their original spatial orientation. Potential fields of application are rapid assessments of endolithic bio- and ichnodiversity and the quantification of the impact of microendoliths on the overall calcium carbonate turnover. The method can be applied after CLSM of the stained microendoliths and retains the opportunity for a subsequent investigation of epoxy casts with SEM. This allows a three-fold approach in studying microendoliths in the context of their microborings, thereby fostering the integration of biological and ichnological aspects of microbial bioerosion.

Bioerosion describes the process of active erosion of hard substrates induced by the activity of living organisms. Beside numerous marine macroscopic bioeroding organisms such as sponges, annelids or bivalves, there is an astonishing 'hidden diversity' of microscopic bioeroding organisms which produce minute tunnels and chambers, for example in calcareous shells and skeletons of other marine organisms. These so-called microendoliths belong to bacteria, microalgae, foraminiferans, or fungi. Due to their lifestyle hidden inside the hard substrate, scientific investigation is often laborious and involves complex preparation techniques, electron microscopy, or even nano-computed tomography. Photo-autotrophic microendoliths (eg cyanobacteria and algae) have been studied with fluorescence microscopy using autofluorescence properties, for example of their chloroplasts. However, microendoliths of aphotic depths, mostly of fungal origin, do not show autofluorescence. With the present study we test different fluorescent dyes staining the microbioeroders 'in situ', that is, inside their microscopic tunnels, and visualise them using three-dimensional confocal laser scanning microscopy (CLSM). Very good results have been obtained with the dye Sybr Green I that stains DNA molecules and thereby the cell nuclei of the microendoliths. This method can be used, for instance, to measure the infestation rate of a given substrate by discriminating between abandoned microborings and those still inhabited by microendoliths. Another approach that was successfully tested in the course of the present study was the infiltration of the cleaned microborings with resin that was previously mixed with the fluorescent dye Safranin-O. The datasets obtained with the CLSM were used to reconstruct 3D-surface models of the microborings of three different microendoliths. Such models can be used to analyse the original spatial arrangement inside the hard substrate and to measure exact volumes. The resulting possibility to make exact quantifications is of high value for future investigations that focus on the role and proportion of microbioerosion in the (re)cycling of marine carbonates.

High-precision targeting workflow for volume electron microscopy.

Cells are 3D objects. Therefore, volume EM (vEM) is often crucial for correct interpretation of ultrastructural data. Today, scanning EM (SEM) methods such as focused ion beam (FIB)-SEM are frequently used for vEM analyses. While they allow automated data acquisition, precise targeting of volumes of interest within a large sample remains challenging. Here, we provide a workflow to target FIB-SEM acquisition of fluorescently labeled cells or subcellular structures with micrometer precision. The strategy relies on fluorescence preservation during sample preparation and targeted trimming guided by confocal maps of the fluorescence signal in the resin block. Laser branding is used to create landmarks on the block surface to position the FIB-SEM acquisition. Using this method, we acquired volumes of specific single cells within large tissues such as 3D cultures of mouse mammary gland organoids, tracheal terminal cells in Drosophila melanogaster larvae, and ovarian follicular cells in adult Drosophila, discovering ultrastructural details that could not be appreciated before.

Efeito do acabamento e polimento sobre a topografia superficial de uma resina composta convencional e uma bulk fill / Effect of finishing and polishing on the surface topography of a conventional compound resin and a bulk fill / Efecto de acabado y pulido en la topografía superficial de una resina compuesta convencional y un llenado a granel

Introdução:As atuaisresinas Bulk-Fill apresentamcomovantagens e propriedades: menor tempo clínico, baixa contração de polimerização, maior translucidez, entre outros. Objetivo:Avaliar a topografia superficial de restaurações com diferentes tipos de resina composta antes e após submetê-las a diferentes sistemas de acabamento e polimento. Metodologia:Confecção de100 corpos de prova, sendo 50 de cada composto resinoso, apresentando cinco grupos (n=10) sorteados de forma aleatória, de acordo com o sistema de acabamento e polimento empregado. Foram utilizadas as resinas Filtek Z350 XT e a Bulk Fill Filtek One Bulk Fill e os sistemas de acabamento e polimento: Discos de Lixa Sof-Lex Pop On Kit;Discos Diamantado Espiral Sof-Lex; Ponta Enhance e Broca Carbide Multilaminada nº 0283F, que foram comparados com superfície deixada pela Tira de Poliester (Controle Negativo). Após 7 dias de armazenamento em água destilada, os corpos de provaforam submetidos aoMicroscópio Eletrônico de Varredura.Resultados:Os corpos de provada resina Bulk Fill tiveram como melhor resultado o sistema de acabamento e polimentocom broca carbidee resultado não satisfatório com discodiamantado espiral Sof-lex, enquanto a ponta Enhance e lixa Sof-lex Pop Ontiveram resultados semelhantes. Os corpos de prova da resina Filtek Z350 XT tiveram melhor aspecto visual de lisura no grupo controle negativoe pior resultado no grupo com disco diamantado espiral Sof-lex. Conclusões:Dessa forma, o sistema que apresentou melhor resultado foi a broca Carbidena resina Bulk Fill, enquanto a Filtek Z350 XT apresentou-se melhor no grupo controle. Porém, ainda são necessáriosestudos para se chegar em um protocolo de acabamento e polimento mais eficiente (AU).

Introduction:The current Bulk-Fill resins have the following advantages and properties: less clinical time, low polymerization shrinkage, greater translucency, among others.Objective:Toevaluate the topography of restorations with different types of composite resin before and after submitting them to different finishing and polishing systems.Methodology:Manufacture of 100 specimens, 50 of each resin compound, presenting five groups (n = 10) drawn at random, according to the finishing and polishing system employed. Filtek Z350 XT resins and Bulk Fill Filtek One Bulk Fill andfinishing and polishing systems were used: Sof-Lex Pop On Kit Sanding Discs; Diamond Spiral Sof-Lex Discs; Tip Enhance and Multilaminated Carbide Drill nº 0283F, which were compared with the surface left by the Polyester Strip (Negative Control). After 7 days of storage in distilled water, the specimens were submitted to a Scanning Electron Microscope.Results:The Bulk Fill resin specimens had the best result of the finishing and polishing system with carbide drill and unsatisfactory result with Sof-lex spiral diamond disc, while the Enhance tip and Sof-lex Pop On sandpaper had similar results. The Filtek Z350 XT resin specimens had a better visual aspect of smoothness in the negative control group and a worse result in the group with Sof-lex spiral diamond wheel.Conclusions:Thus, the system that showed the best result was the Carbide drill in Bulk Fill resin, while Filtek Z350 XT was better in the control group. However, studies are still needed to arrive at a more efficient finishing and polishing protocol (AU).

Introducción:Las actuales resinas Bulk-Fill tienen las siguientes ventajas y propiedades: menor tiempo clínico, baja contracción de polimerización, mayor translucidez, entre otras.Objetivo: Evaluar la topografía de restauraciones con diferentes tipos de resina compuesta antes y después de someterlas a diferentes sistemas de acabado y pulido.Metodología: Fabricación de 100 probetas, 50 de cada resina compuesta, presentando cinco grupos (n = 10) extraídos al azar, según el sistema de acabado y pulido empleado. Se utilizaron resinas Filtek Z350 XT y Bulk Fill Filtek One Bulk Fill y sistemas de acabado y pulido: Discos de lijado Sof-Lex Pop On Kit; Discos Sof-Lex en espiral de diamante; Broca Realce de Punta y Carburo Multilaminado nº 0283F, que fueron comparadas con la superficie dejada por la Tira de Poliéster (Control Negativo). Después de 7 días de almacenamiento en agua destilada, lasmuestras se sometieron a un Las muestras de resina Bulk Fill tuvieron el mejor resultado del sistema de acabado y pulido con broca de carburo y un resultado insatisfactorio con el disco de diamante en espiral Sof-lex, mientras que la punta Enhance y el papel de lija Sof-lex Pop On tuvieron resultados similares. Las muestras de resina Filtek Z350 XT tuvieron un mejor aspecto visual de suavidad en el grupo de control negativo y un peor resultado en el grupo con disco de diamante en espiral Sof-lex.Conclusiones: Así, el sistema que mejor resultado mostró fue la broca Carbide en resina Bulk Fill, mientras que Filtek Z350 XT fue mejor en el grupo control. Sin embargo, aún se necesitan estudios para llegar a un protocolo de acabado y pulido más eficiente (AU).

Tuneable in-situ nanoCT workflow using FIB/SEM.

Inspired by the standard computed tomography, a new method of 3D X-ray imaging embedded in FIB-SEM microscope is proposed. The unique combination of TEM-like specimen stage enabling in lens STEM detection (referred to as CompuStage), nanomanipulator (referred to as EasyLift) facilitating in-situ sample transfer from bulk sample to TEM-like stage and pixelated in-situ Timepix X-ray detector in Helios G4 FX FIB-SEM system offers an unprecedented workflow. Motivated by common circular CT scan known from microCT world, the object under study is placed on CompuStage rod which enables two possible rotation (in TEM/SEM terminology called tilt) movements - α-tilt - rotation of the CompuStage rod around its axis, and ß-tilt - rotation around axis perpendicular to CompuStage rod. ß-tilt rotation enables a circular movement of the sample while α-tilt sets the correct position of sample with respect to target and detector. Thin metal lamella of suitable material welded to EasyLift manipulator needle is used as an X-ray target. The final target-sample geometry - position, distance - can be fine-tuned using position control of CompuStage and EasyLift and in-situ monitored by SEM. Both sample and target can also be easily prepared in-situ. Radiographs are recorded by Timepix detector with inherent noise-free operation and energy filtration. For the 3D reconstruction standard microCT reconstruction algorithm is used with the procedure adjusted for the format and quality of nanoCT images. The experiments were carried out on Helios G4 FX DualBeam using titanium and tungsten targets and various semiconductor samples. The ultimate resolution of the proposed method in orders of tens of nanometers was achieved both by the possibility of close target to sample positioning and of adjustment of primary beam energy down to low energies reducing the interaction volume in the target. Since the lower energy radiation is well suited for life-science, the method was also tested on several bio-samples using silver target. The silver target, thanks to its massive low energy Lα line, allowed to distinguish subtle structures in the resin embedded stained mouse brain and also to observe and reconstruct canaliculi in the mouse bone (earlier reported by Dierolf et al. 2010, Nature 467 436).

Topography of Primary Molar Pulp Chamber Floor: A Scanning Electron Microscopy and Micro-Computed Tomography Analysis

ABSTRACT Objective: To determine in vitro the frequency, shape, type, diameter, and patency of accessory canals in the primary molars pulp chamber floor. Material and Methods: Sixteen healthy primary molars were evaluated by micro-computed tomography and scanning electron microscopy. Descriptive analyses of the frequency, shape (round, oval, or irregular), type (blind, true, or hidden), patency and diameter of the accessory canals were performed. Results: Half of the teeth presented accessory canals, 62.5% of which were located in the upper molars and 37.5% in the lower molars. The most frequent shape was irregular. In three-dimensional analysis, blind accessory canals (12.5%) and with patency (18.7%) of the teeth were observed. The average accessory canal diameter was 51.97 µm (± 26.03 µm). Conclusion: Upper molars showed a higher frequency of accessory canals with larger diameters. The irregular shape was the most frequent. 18.7% of accessory channels showed patency.

Characteristics of Starch Extracted from the Stem of Pineapple Plant (Ananas comosus) - an Agro Waste from Pineapple Farms

Abstract The present study focused on the use of pineapple plant stem, which is an agro-waste, for the production of starch (11.08 % ± 0.77). Characters were studied using X-ray diffraction, nuclear magnetic resonance spectroscopy (NMR), fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), differential scanning calorimetry (DSC) and rheological methods. The granular size of stem starch was comparatively smaller than corn starch granules. The X-ray diffraction data revealed that stem starch has an A-type crystal structure. The molecular structure was similar to those obtained for native starches, which is confirmed by NMR and FTIR. The gelatinization temperature was observed to be higher than corn starch and rheological studies revealed; stem starch is more viscous than corn starch. The purity analysis showed that the harmful heavy metals were in negligible quantity and the tested pesticides were absent. This could make this a good source of starch for food industries. Results revealed that this agro-waste has a high potential for the production of good quality starch.

Short Communication: An Updated Design to Implement Artificial Neuron Synaptic Behaviors in One Device with a Control Gate.

BACKGROUND: As a key component in artificial intelligence computing, a transistor design is updated here as a potential alternative candidate for artificial synaptic behavior implementation. However, further updates are needed to better control artificial synaptic behavior. Here, an updated channel-electrode transistor design is proposed as an artificial synapse device; this structure is different from previously published designs by other groups. METHODS: A semiconductor characterization system was used in order to simulate the artificial synaptic behavior and a scanning electron microscope was used to characterize the device structure. RESULTS: It was found that the electrode added to the transistor channel had a strong impact on the representative transmission behavior of such artificial synaptic devices, such as excitatory postsynaptic current (EPSC) and the paired-pulse facilitation (PPF) index. CONCLUSION: These behaviors were tuned effectively and the impact of the channel electrode is explained by the combined effects of the joint channel electrode and conventional gate. The voltage dependence of such oxide devices suggests more capability to emulate various synaptic behaviors for numerous medical and non-medical applications. This is extremely helpful for future neuromorphic computational system implementation.

Imaging Plant Cells by High-Pressure Freezing and Serial block-face scanning electron microscopy.

This chapter describes methods to enhanced contrast of plant material processed by high-pressure freezing and freeze substitution for improved visualization by serial block-face scanning electron microscopy (SBEM). The contrast enhancing steps are based on a protocol involving the sequential incubation of samples in heavy metals and sodium thiocarbohydrazide (OTO staining). We also describe the pipeline for imaging plant tissues in a commercial SBEM system (Gatan 3View®) and routines for the image analysis and three-dimensional reconstructions using open-source and commercial software packages.

Imaging dataset of fresh hydrous plants obtained by field-emission scanning electron microscopy conducted using a protective NanoSuit.

Although scanning electron microscopy (SEM) can generate high-resolution images of nanosized objects, it requires a high vacuum to do so, which precludes direct observations of living organisms and often produces unwanted structural changes. It has previously been reported that a simple surface modification gives rise to a nanoscale layer, termed the "NanoSuit", which can keep small animals alive under the high vacuum required for field-emission scanning electron microscopy (FE-SEM). We have previously applied this technique to plants, and successfully observed healthy petals in a fully hydrated state using SEM. The flower petals protected with the NanoSuit appeared intact, although we still lack a fundamental understanding of the images of other plants observed using FE-SEM. This report presents and evaluates a rich set of images, acquired using the NanoSuit, for a taxonomically diverse set of plant species. This dataset of images allows the surface features of various plants to be analyzed and thus provides a further complementary morphological profile. Image data can be accessed and viewed through Figshare (https://doi.org/10.6084/m9.figshare.c.4446026.v1).

Electron Microscopy Techniques Applied to Symbiont-Harboring Trypanosomatids: The Association of the Bacterium with Host Organelles.

In this chapter we describe different electron microscopy techniques such as freeze fracture, deep etching, and three-dimensional reconstruction, obtained by electron tomography or focused ion beam scanning electron microscopy (FIB-SEM), combined with quick-freezing methods in order to reveal aspects of the cell structure in trypanosomatids. For this purpose, we chose protists that evolve in a mutualistic way with a symbiotic bacterium. Such cells represent excellent models to study the positioning and distribution of organelles, since the symbiotic bacterium interacts with different organelles of the host trypanosomatid. We demonstrate that the employment of such techniques can show the proximity and even the interaction of the symbiotic bacterium with different structures of the protist host, such as the nucleus and the glycosomes. In addition, the quick-freezing approach can reveal new aspects of the gram-negative bacterial envelope, such as the presence of a greatly reduced cell wall between the two membrane units.

Transmission Fourier Transform Infrared Spectroscopic Imaging, Mapping, and Synchrotron Scanning Microscopy with Zinc Sulfide Hemispheres on Living Mammalian Cells at Sub-Cellular Resolution.

Fourier transform infrared (FT-IR) spectroscopic imaging and microscopy of single living cells are established label-free technique for the study of cell biology. The constant driver to improve the spatial resolution of the technique is due to the diffraction limit given by infrared (IR) wavelength making subcellular study challenging. Recently, we have reported, with the use of a prototype zinc sulfide (ZnS) transmission cell made of two hemispheres, that the spatial resolution is improved by the factor of the refractive index of ZnS, achieving a λ/2.7 spatial resolution using the synchrotron-IR microscopy with a 36× objective with numerical aperture of 0.5. To refine and to demonstrate that the ZnS hemisphere transmission device can be translated to standard bench-top FT-IR imaging systems, we have, in this work, modified the device to achieve a more precise path length, which has improved the spectral quality of the living cells, and showed for the first time that the device can be applied to study live cells with three different bench-top FT-IR imaging systems. We applied focal plane array (FPA) imaging, linear array, and a synchrotron radiation single-point scanning method and demonstrated that in all cases, subcellular details of individual living cells can be obtained. Results have shown that imaging with the FPA detector can measure the largest area in a given time, while measurements from the scanning methods produced a smoother image. Synchrotron radiation single-point mapping produced the best quality image and has the flexibility to introduce over sampling to produce images of cells with great details, but it is time consuming in scanning mode. In summary, this work has demonstrated that the ZnS hemispheres can be applied in all three spectroscopic approaches to improve the spatial resolution without any modification to the existing microscopes.