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1.
Nat Commun ; 11(1): 996, 2020 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-32081905

RESUMO

Serial X-ray crystallography at free-electron lasers allows to solve biomolecular structures from sub-micron-sized crystals. However, beam time at these facilities is scarce, and involved sample delivery techniques are required. On the other hand, rotation electron diffraction (MicroED) has shown great potential as an alternative means for protein nano-crystallography. Here, we present a method for serial electron diffraction of protein nanocrystals combining the benefits of both approaches. In a scanning transmission electron microscope, crystals randomly dispersed on a sample grid are automatically mapped, and a diffraction pattern at fixed orientation is recorded from each at a high acquisition rate. Dose fractionation ensures minimal radiation damage effects. We demonstrate the method by solving the structure of granulovirus occlusion bodies and lysozyme to resolutions of 1.55 Å and 1.80 Å, respectively. Our method promises to provide rapid structure determination for many classes of materials with minimal sample consumption, using readily available instrumentation.


Assuntos
Cristalografia/métodos , Proteínas/química , Microscopia Eletrônica de Transmissão e Varredura , Modelos Moleculares , Muramidase/química , Muramidase/ultraestrutura , Nanopartículas/química , Nanopartículas/ultraestrutura , Proteínas de Matriz de Corpos de Inclusão/química , Proteínas de Matriz de Corpos de Inclusão/ultraestrutura , Tamanho da Partícula , Conformação Proteica , Proteínas/ultraestrutura
2.
Chemosphere ; 239: 124773, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31518919

RESUMO

Hyporheic zone (HZ) sediments in river systems are often contaminated with heavy metals as a legacy of natural processes and anthropogenic activities. The geochemical behaviors of heavy metals in the HZ sediments at the laboratory scale have been extensively studied. However, the watershed-scale distributions of heavy metals in the HZ sediments and the processes controlling their distributions have not been well studied. Here, we report a watershed-scale study of heavy metals (i.e., Cr, Ni, Cu, Zn, Cd, and Pb) distributions in the HZ of the Maozhou River watershed, a heavily polluted area within the Pearl River Delta, southern China. Statistical analysis revealed that the spatial distribution of studied heavy metal concentrations was highly correlated with that of the sediment-associated sulfide at the watershed-scale. Metal extraction analysis and double-spherical aberration-corrected scanning transmission electron microscope imaging (Cs-STEM) further confirmed the strong association of heavy metals with sulfur. These observations demonstrated that the formation of metals-sulfide precipitates was the key process controlling the watershed-scale distributions of heavy metals (especially for Cr, Ni and Zn) in the HZ sediments. Additionally, high permeability of the HZ sediments may prevent Ni, Zn, Cd and Pb accumulation in sediments. Specially, Cu distribution was mainly affected by organic-Cu complexation. In the estuary area, salinity input likely affected the distributions of Ni, Zn and Cd through cation exchange processes. The findings improved our understanding of the distributions of heavy metals and the processes controlling their distributions at the watershed-scale, and have implications for remediating and managing contaminated HZ sediments.


Assuntos
Sedimentos Geológicos/análise , Metais Pesados/análise , Poluentes Químicos da Água/análise , China , Monitoramento Ambiental/métodos , Estuários , Sedimentos Geológicos/química , Metais Pesados/química , Metais Pesados/isolamento & purificação , Microscopia Eletrônica de Transmissão e Varredura , Rios , Salinidade , Sulfetos/análise , Sulfetos/química , Poluentes Químicos da Água/química , Poluentes Químicos da Água/isolamento & purificação
3.
Chemosphere ; 239: 124775, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31521931

RESUMO

Enriched levels of thallium (Tl) in the environment are not only derived from anthropogenic sources but also have potential natural origins owing to Tl-rich sulphide mineralization. However, little is known regarding the geochemical fractionations of Tl in contaminated soils from geogenic sources. This study aims to reveal the Tl geochemical fractionations in different types of soils from a large-scale independent Tl mine in southwestern China, via a modified Institute for Reference Materials and Measurement (IRMM) sequential extraction (four-step) scheme. The results revealed that a large percentage of Tl was related to the labile portions (including reducible, weak-acid-exchangeable, and oxidizable fraction) of the soils (68.8-367 mg kg-1). Further analyses by Scanning Transmission Electron Microscopy-Energy Dispersive X-ray Spectrometer (STEM-EDS) found that Tl mainly existed in the Fe-containing minerals (such as jarosite and hematite) with fine particles (∼1 µm). These results highlight that, apart from the anthropogenically induced Tl pollution, the naturally occurring Tl contamination in soils may also pose significant risks to human health and ecological safety. Owing to the relatively high mobility and bioavailability of Tl in the labile fractions, it is important to understand geochemical fractionations of this element for alleviating Tl pollution and effective management of naturally occurring Tl contaminated soils.


Assuntos
Poluentes do Solo/química , Tálio/química , Fracionamento Químico , China , Monitoramento Ambiental/métodos , Compostos Férricos/química , Humanos , Mercúrio/análise , Microscopia Eletrônica de Transmissão e Varredura , Solo/química , Poluentes do Solo/análise , Espectrometria por Raios X , Sulfatos/química , Sulfetos/análise , Tálio/análise
4.
PLoS One ; 14(12): e0226277, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31860683

RESUMO

Filamentous proteins are responsible for the superior mechanical strength of our cells and tissues. The remarkable mechanical properties of protein filaments are tied to their complex molecular packing structure. However, since these filaments have widths of several to tens of nanometers, it has remained challenging to quantitatively probe their molecular mass density and three-dimensional packing order. Scanning transmission electron microscopy (STEM) is a powerful tool to perform simultaneous mass and morphology measurements on filamentous proteins at high resolution, but its applicability has been greatly limited by the lack of automated image processing methods. Here, we demonstrate a semi-automated tracking algorithm that is capable of analyzing the molecular packing density of intra- and extracellular protein filaments over a broad mass range from STEM images. We prove the wide applicability of the technique by analyzing the mass densities of two cytoskeletal proteins (actin and microtubules) and of the main protein in the extracellular matrix, collagen. The high-throughput and spatial resolution of our approach allow us to quantify the internal packing of these filaments and their polymorphism by correlating mass and morphology information. Moreover, we are able to identify periodic mass variations in collagen fibrils that reveal details of their axially ordered longitudinal self-assembly. STEM-based mass mapping coupled with our tracking algorithm is therefore a powerful technique in the characterization of a wide range of biological and synthetic filaments.


Assuntos
Colágeno/química , Processamento de Imagem Assistida por Computador/métodos , Proteínas dos Microfilamentos/química , Actinas/química , Algoritmos , Animais , Humanos , Microscopia Eletrônica de Transmissão e Varredura , Microtúbulos/química
5.
Micron ; 127: 102759, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31585250

RESUMO

Copper surface after antibacterial test against E. coli was examined in the aspect of corrosion. Results from scanning electron microscope (SEM), grazing incidence X-ray diffractometer (GIXRD) and Raman spectroscopy together confirmed less oxidation on copper surface with the presence of E. coli. The inhibition of the cuprous oxide (Cu2O) layer instead ensured the continuous exposure of copper surface, letting localised corrosion attacks observable and causing a stronger release of copper ions. These phenomena are attributed to the fact that E. coli act as ions reservoirs since high amount of copper accumulation were found by energy dispersive X-ray spectroscopy (EDS).


Assuntos
Cobre/química , Escherichia coli/metabolismo , Corrosão , Microscopia Eletrônica de Transmissão e Varredura , Oxirredução , Propriedades de Superfície
6.
Nanoscale ; 11(36): 16868-16878, 2019 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-31482911

RESUMO

Ferritin is a protein that regulates the iron ions in humans by storing them in the form of iron oxides. Despite extensive efforts to understand the ferritin iron oxide structures, it is still not clear how ferritin proteins with a distinct light (L) and heavy (H) chain subunit ratio impact the biomineralization process. In situ graphene liquid cell-transmission electron microscopy (GLC-TEM) provides an indispensable platform to study the atomic structure of ferritin mineral cores in their native liquid environment. In this study, we report differences in the iron oxide formation in human spleen ferritins (HSFs) and human heart ferritins (HHFs) using in situ GLC-TEM. Scanning transmission electron microscopy (STEM) along with selected area electron diffraction (SAED) of the mineral core and electron energy loss spectroscopy (EELS) analyses enabled the visualization of morphologies, crystal structures and the chemistry of iron oxide cores in HSFs and HHFs. Our study revealed the presence of metastable ferrihydrite (5Fe2O3·9H2O) as a dominant phase in hydrated HSFs and HHFs, while a stable hematite (α-Fe2O3) phase predominated in non-hydrated HSFs and HHFs. In addition, a higher Fe3+/Fe2+ ratio was found in HHFs in comparison with HSFs. This study provides new understanding on iron-oxide phases that exist in hydrated ferritin proteins from different human organs. Such new insights are needed to map ferritin biomineralization pathways and possible correlations with various iron-related disorders in humans.


Assuntos
Compostos Férricos/metabolismo , Microscopia Eletrônica de Transmissão e Varredura , Miocárdio , Baço , Ferritinas , Grafite , Humanos , Miocárdio/metabolismo , Miocárdio/ultraestrutura , Baço/metabolismo , Baço/ultraestrutura
7.
Methods Cell Biol ; 152: 197-215, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31326021

RESUMO

Electron cryo-tomography using the scanning transmission modality (STEM) enables 3D reconstruction of unstained, vitrified specimens as thick as 1µm or more. Contrast is related to mass/thickness and atomic number, providing quantifiable chemical characterization and mass mapping of intact prokaryotic and eukaryotic cells. Energy dispersive X-ray spectroscopy by STEM provides a simple, on-the-spot chemical identification of the elemental composition in sub-cellular organic bodies or mineral deposits. This chapter provides basic background and practical information for performing cryo-STEM tomography on vitrified biological cells.


Assuntos
Biologia/métodos , Microscopia Crioeletrônica/métodos , Microscopia Eletrônica de Transmissão e Varredura/métodos , Células Eucarióticas/fisiologia , Imageamento Tridimensional/métodos , Células Procarióticas/fisiologia , Espectrometria por Raios X/métodos
8.
Environ Pollut ; 252(Pt A): 289-295, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31158657

RESUMO

The Athabasca Oil Sands Region (AOSR) in Alberta, Canada, is an important source of atmospheric pollutants, such as aerosols, that have repercussions on both the climate and human health. We show that the mean freezing temperature of snow-borne particles from AOSR was elevated (-7.1 ±â€¯1.8 °C), higher than mineral dust which freezes at ∼ -15 °C and is recognized as one of the most relevant ice nuclei globally. Ice nucleation of nanosized snow samples indicated an elevated freezing ability (-11.6 ±â€¯2.0 °C), which was statistically much higher than snow-borne particles from downtown Montreal. AOSR snow had a higher concentration (∼2 orders of magnitude) of >100 nm particles than Montreal. Triple quadrupole ICP-(QQQ)-MS/MS analysis of AOSR and Montreal snow demonstrated that most concentrations of metals, including those identified as emerging nanoparticulate contaminants, were much more elevated in AOSR in contrast to Montreal: 34.1, 34.1, 16.6, 5.8, 0.3, 0.1, and 9.4 mg/m3 for Cr, Ni, Cu, As, Se, Cd, and Pb respectively, in AOSR and 1.3, 0.3, 2.0, <0.03, 0.1, 0.03, and 1.2 mg/m3 in Montreal snow. High-resolution Scanning Transmission Electron Microscopy/Energy-dispersive X-ray Spectroscopy (STEM-EDS) imaging provided evidence for various anthropogenic nano-materials, including carbon nanotubes resembling structures, in AOSR snow up to 7-25 km away from major oil sands upgrading facilities. In summary, particles characterized as coming from oil sands are more efficient at ice nucleation. We discuss the potential impacts of AOSR emissions on atmospheric and microphysical processes (ice nucleation and precipitation) both locally and regionally.


Assuntos
Poluição do Ar/análise , Monitoramento Ambiental/métodos , Nanopartículas Metálicas/análise , Nanotubos de Carbono/análise , Neve/química , Aerossóis/análise , Alberta , Poeira/análise , Gelo , Microscopia Eletrônica de Transmissão e Varredura , Campos de Petróleo e Gás , Tamanho da Partícula , Espectrometria por Raios X , Espectrometria de Massas em Tandem
9.
Vis Neurosci ; 36: E004, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-31199211

RESUMO

There are more than 30 distinct types of mammalian retinal ganglion cells, each sensitive to different features of the visual environment. In rabbit retina, they can be grouped into four classes according to their morphology and stratification of their dendrites in the inner plexiform layer (IPL). The goal of this study was to describe the synaptic inputs to one type of Class IV ganglion cell, the third member of the sparsely branched Class IV cells (SB3). One cell of this type was partially reconstructed in a retinal connectome developed using automated transmission electron microscopy (ATEM). It had slender, relatively straight dendrites that ramify in the sublamina a of the IPL. The dendrites of the SB3 cell were always postsynaptic in the IPL, supporting its identity as a ganglion cell. It received 29% of its input from bipolar cells, a value in the middle of the range for rabbit retinal ganglion cells studied previously. The SB3 cell typically received only one synapse per bipolar cell from multiple types of presumed OFF bipolar cells; reciprocal synapses from amacrine cells at the dyad synapses were infrequent. In a few instances, the bipolar cells presynaptic to the SB3 ganglion cell also provided input to an amacrine cell presynaptic to the ganglion cell. There was apparently no crossover inhibition from narrow-field ON amacrine cells. Most of the amacrine cell inputs were from axons and dendrites of GABAergic amacrine cells, likely providing inhibitory input from outside the classical receptive field.


Assuntos
Células Amácrinas/ultraestrutura , Células Bipolares da Retina/ultraestrutura , Células Ganglionares da Retina/ultraestrutura , Sinapses/ultraestrutura , Animais , Conectoma , Feminino , Neurônios GABAérgicos/ultraestrutura , Microscopia Eletrônica de Transmissão e Varredura , Microscopia Eletrônica de Transmissão , Coelhos
10.
Int J Low Extrem Wounds ; 18(3): 323-335, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31140339

RESUMO

Tissue regeneration has become a promising strategy for repairing damaged skin tissues. Among the hydrogels for tissue regeneration applications, topical hydrogels have demonstrated great potential for use as 3D-scaffolds in the burn wound healing process. Currently, no report has been published specifically on icariin-loaded polyvinyl alcohol (PVA)/agar hydrogel on full-thickness burn wounds. In the present study, burn tissue regeneration based on biomimetic hydrogel scaffolds was used for repairing damaged extracellular matrix. Furthermore, a skin burn model was developed in rats, and the icariin-loaded PVA/agar hydrogels were implanted into the damaged portions. The regeneration of the damaged tissues with the help of the icariin-loaded hydrogel group exhibited new translucent skin tissues and repaired extracellular matrix, indicating that the hydrogel can enhance the wound healing process. Moreover, characterization studies such as X-ray diffraction, Fourier-transformed infrared spectroscopy, and differential scanning calorimetry reported the extent of compatibility between icariin and its polymers. Results of the field emission scanning electron microscopy images revealed the extent of the spread of icariin within the polymer-based hydrogel. Furthermore, the wound healing potential, confirmed by histopathological and histochemical findings at the end of 21 days, revealed the visual evidence for the biomimetic property of icariin-loaded PVA/agar hydrogel scaffolds with the extracellular matrix for tissue regeneration.


Assuntos
Queimaduras , Flavonoides/farmacologia , Regeneração/efeitos dos fármacos , Lesões dos Tecidos Moles , Cicatrização , Animais , Queimaduras/patologia , Queimaduras/terapia , Varredura Diferencial de Calorimetria/métodos , Medicamentos de Ervas Chinesas , Excipientes/farmacologia , Microscopia Eletrônica de Transmissão e Varredura/métodos , Modelos Animais , Álcool de Polivinil/farmacologia , Ratos , Lesões dos Tecidos Moles/diagnóstico por imagem , Lesões dos Tecidos Moles/patologia , Lesões dos Tecidos Moles/terapia , Tecidos Suporte , Resultado do Tratamento , Cicatrização/efeitos dos fármacos , Cicatrização/fisiologia
11.
Spectrochim Acta A Mol Biomol Spectrosc ; 220: 117139, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31146209

RESUMO

The determination of neomycin sulfate was made using photoluminescent amino-functionalized graphene quantum dots (obtained from hydro-exfoliation of a mixture of citric acid and glutathione). From the several ions tested, Fe3+ was the best mediator to enable an off/on photoluminescence effect used for quantification. The mediation of Fe3+ was found to be crucial as it is responsible for the photoluminescence quenching effect, due to the interaction with quantum dots surface, also having large affinity towards neomycin that removes Fe3+ from the surface of GQDs, consequently, promoting restoration of the original nanomaterial photoluminescence. Such signal restoration was proportional to the neomycin sulfate concentration added. The linearized analytical response covered three orders of magnitude (10-7 to 10-5 mol L-1). The proposed method is an alternative to those requiring labor-intensive procedures for chemical the derivatization of neomycin (due to the lack of chromophore groups in aminoglycosides). The method was successfully tested in the analysis of rubella vaccine containing trace residues of neomycin and in pharmaceutical compositions containing neomycin sulfate after solid phase extraction using an aminoglycoside imprinted polymer to improve selectivity in determinations.


Assuntos
Neomicina/análise , Pontos Quânticos/química , Vacina contra Rubéola/análise , Aminoglicosídeos/química , Glutationa/química , Grafite/química , Ferro/química , Limite de Detecção , Medições Luminescentes , Microscopia Eletrônica de Transmissão e Varredura , Impressão Molecular , Sondas Moleculares/química , Espectroscopia Fotoeletrônica , Extração em Fase Sólida/instrumentação , Análise Espectral Raman , Temperatura
12.
Ultramicroscopy ; 202: 44-50, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30953993

RESUMO

In sample preparation of biological samples for electron microscopy, many types of embedding media are widely used. Unfortunately, none of them is perfectly resistant to beam induced damage. The article is focused on mass loss measuring of pure epoxy resin EMbed 812 that replaced Epon - the most widely used embedding resin for biological electron microscopy, in a form of ultrathin sections with thicknesses ranging from 30 to 100 nm. The STEM imaging was performed in a quantitative way which allowed us to estimate the mass loss directly up to the total dose of 3000 e-/nm2. For data acquisition we used SEM equipped with a commercial STEM detector working at a relatively low acceleration voltage of 30 kV. In this study we estimated the influence of various factors which can affect the endurance of the epoxy resin EMbed 812 ultrathin sections under an electron beam, such as the sample aging, differences between storing the samples in forms of ultrathin sections and whole blocks, ultrathin sections thicknesses, temperature of the sample, probe current, and one or two-sided carbon coating of ultrathin sections. The aim of this work is to investigate beam induced mass loss at electron energies of SEM and find out how to reduce the mass loss.


Assuntos
Resinas Epóxi/química , Microscopia Eletrônica de Transmissão e Varredura/métodos , Elétrons
13.
Food Chem Toxicol ; 127: 89-100, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30849403

RESUMO

The aim of this study was to evaluate cytotoxicity (WST-1 assay), LDH release (LDH assay) and genotoxicity (Comet assay) of three engineered TiO2-NPs with different shapes (bipyramids, rods, platelets) in comparison with two commercial TiO2-NPs (P25, food grade). After NPs characterization (SEM/T-SEM and DLS), biological effects of NPs were assessed on BEAS-2B cells in presence/absence of light. The cellular uptake of NPs was analyzed using Raman spectroscopy. The cytotoxic effects were mostly slight. After light exposure, the largest cytotoxicity (WST-1 assay) was observed for rods; P25, bipyramids and platelets showed a similar effect; no effect was induced by food grade. No LDH release was detected, confirming the low effect on plasma membrane. Food grade and platelets induced direct genotoxicity while P25, food grade and platelets caused oxidative DNA damage. No genotoxic or oxidative damage was induced by bipyramids and rods. Biological effects were overall lower in darkness than after light exposure. Considering that only food grade, P25 and platelets (more agglomerated) were internalized by cells, the uptake resulted correlated with genotoxicity. In conclusion, cytotoxicity of NPs was low and affected by shape and light exposure, while genotoxicity was influenced by cellular-uptake and aggregation tendency.


Assuntos
Brônquios/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Células Epiteliais/efeitos dos fármacos , Nanopartículas Metálicas/toxicidade , Titânio/toxicidade , Brônquios/citologia , Brônquios/enzimologia , Linhagem Celular , Dano ao DNA , Células Epiteliais/enzimologia , Humanos , L-Lactato Desidrogenase/metabolismo , Microscopia Eletrônica de Transmissão e Varredura , Estresse Oxidativo/efeitos dos fármacos , Tamanho da Partícula , Análise Espectral Raman/métodos
14.
Micron ; 119: 54-63, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30660856

RESUMO

In the use of solution-based 3D nanoarchitectures for optics, drug delivery, and cancer treatment, the precise nanoparticle architecture morphologies, architecture sizes, interparticle distances, and the assembly stability are all critical to their functionality. 3D nanoparticle architectures in solution are difficult to characterize, as few techniques can provide individualized information on interparticle spacing (defined by linkage molecule), nanoparticle assembly size, morphology, and identification of false aggregation. Bulk characterization techniques, including small angle x-ray scattering, can provide architecture sizes, though they are unable to precisely measure differences within interparticle spacings for individual architectures and can falsely measure assemblies caused by non-linkage grouped nanoparticles. Two solution-based characterization techniques were used to determine which assembly type and linkage length would produce the fastest assembly rate for large DNA-directed gold nanoparticle assemblies. In-situ liquid-cell scanning transmission electron microscopy (LC-STEM), measured interparticle spacings between DNA-functionalized nanoparticles, and fluorescence correlation spectroscopy provided the bulk volume fraction of large and small assemblies for nanoparticle architectures that were assembled using two different types: (1) the hybrid assemblies join two complementary single-stranded DNA linkages, and (2) the bridged assemblies are comprised of single-stranded DNA (bridging component) that is double the length of two different complementary single-stranded DNA-functionalized gold nanoparticles. Assembly times were tested at 24-hrs intervals over 3 days. Statistics derived from the in-situ LC-STEM images provided data for interparticle distance measurements, which identified the fraction of nanoparticles within the images acquired that were at the expected double-stranded DNA-binding distance of the linkages (varied in three distances for each of the two different architectures). In general, longer linkage lengths assembled in the shortest amount of time. The bridged assemblies formed fewer large architectures at 24-hrs but ultimately assembled a greater fraction of nanoparticles, which was due to the longer functionalized DNA lengths for individual nanoparticles. Fluorescence correlation spectroscopy provided a bulk average of the gold nanoparticle assembly sizes over time, which supported the conclusions drawn from the in-situ LC-STEM data. The microscopy provided sub-2 nm precision in the interparticle distances between gold nanoparticles in a solution environment. This coupled microscopy and spectroscopy characterization approach can provide more detailed information than bulk characterization methods.


Assuntos
DNA de Cadeia Simples/metabolismo , Ouro/metabolismo , Nanopartículas Metálicas/ultraestrutura , Microscopia Eletrônica de Transmissão e Varredura/métodos , Espectrometria de Fluorescência/métodos , Cinética , Nanopartículas Metálicas/química , Fatores de Tempo
15.
PLoS One ; 14(1): e0210339, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30699142

RESUMO

The bacterium Sporosarcina pasteurii (SP) is known for its ability to cause the phenomenon of microbially induced calcium carbonate precipitation (MICP). We explored bacterial participation in the initial stages of the MICP process at the cellular length scale under two different growth environments (a) liquid culture (b) MICP in a soft agar (0.5%) column. In the liquid culture, ex-situ imaging of the cellular environment indicated that S. pasteurii was facilitating nucleation of nanoscale crystals of calcium carbonate on bacterial cell surface and its growth via ureolysis. During the same period, the meso-scale environment (bulk medium) was found to have overgrown calcium carbonate crystals. The effect of media components (urea, CaCl2), presence of live and dead in the growth medium were explored. The agar column method allows for in-situ visualization of the phenomena, and using this platform, we found conclusive evidence of the bacterial cell surface facilitating formation of nanoscale crystals in the microenvironment. Here also the bulk environment or the meso-scale environment was found to possess overgrown calcium carbonate crystals. Extensive elemental analysis using Energy dispersive X-ray spectroscopy (EDS) and X-ray powder diffraction (XRD), confirmed that the crystals to be calcium carbonate, and two different polymorphs (calcite and vaterite) were identified. Active participation of S. pasteurii cell surface as the site of calcium carbonate precipitation has been shown using EDS elemental mapping with Scanning transmission electron microscopy (STEM) and scanning electron microscopy (SEM).


Assuntos
Carbonato de Cálcio/química , Carbonato de Cálcio/metabolismo , Sporosarcina/metabolismo , Sporosarcina/ultraestrutura , Biomineralização , Membrana Celular/química , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Cristalização , Meios de Cultura , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão e Varredura , Nanoestruturas/química , Nanoestruturas/ultraestrutura , Difração de Pó , Espectrometria por Raios X , Sporosarcina/crescimento & desenvolvimento
16.
Chembiochem ; 20(6): 822-830, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30501011

RESUMO

Staining compounds containing heavy elements (electron dyes) can facilitate the visualization of DNA and related biomolecules by using TEM. However, research into the synthesis and utilization of alternative electron dyes has been limited. Here, we report the synthesis of a novel DNA intercalator molecule, bis-acridine uranyl (BAU). NMR spectroscopy and MS confirmed the validity of the synthetic strategy and gel electrophoresis verified the binding of BAU to DNA. For TEM imaging of DNA, two-dimensional DNA origami nanostructures were used as a robust microscopy test object. By using scanning transmission electron microscopy (STEM) imaging, which is favored over conventional wide-field TEM for improved contrast, and therefore, quantitative image analysis, it is found that the synthesized BAU intercalator can render DNA visible, even at the single-molecule scale. For comparison, other staining compounds with a purported affinity towards DNA, such as dichloroplatinum, cisplatin, osmium tetroxide, and uranyl acetate, have been evaluated. The STEM contrast is discussed in terms of the DNA-dye association constants, number of dye molecules bound per base pair, and the electron-scattering capacity of the metal-containing ligands. These findings pave the way for the future development of electron dyes with specific DNA-binding motifs for high-resolution TEM imaging.


Assuntos
Acridinas/química , Complexos de Coordenação/química , DNA/química , Substâncias Intercalantes/química , Imagem Individual de Molécula/métodos , Acridinas/síntese química , Complexos de Coordenação/síntese química , Substâncias Intercalantes/síntese química , Microscopia Eletrônica de Transmissão e Varredura/métodos , Conformação de Ácido Nucleico , Urânio/química
17.
Micron ; 116: 30-35, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30265881

RESUMO

Liquid-cell TEM has enabled an interdisciplinary community of scientists to carry out atomic- / nano-scale studies of solid/liquid interfaces. Nevertheless, the restricted resolution of TEM in liquid media and the necessity to reduce the electron dose to avoid harmful radiolytic effects induced by the beam have limited the use of high resolution imaging to study the atomic structure of nanomaterials in liquid. Here we show that STEM nanodiffraction can be exploited in liquid-cell TEM experiments to overcome these two limitations. We evidence that this technique allows quick analysis of the structure of single gold nanoparticles whatever their zone axis orientation, which substantially increases the percentage of analysable nanostructures with respect to HRTEM investigations. Moreover, STEM nanodiffraction can also be used in very low dose conditions. The electron dose irradiating the analyzed nanostructures during data acquisition can be reduced by almost four orders of magnitude compared to conventional HRTEM analysis. Finally, dynamical analyses in reciprocal space are used to provide new insights into the shape-dependent rotation of nanocrystals in the liquid-cell.


Assuntos
Ouro , Nanopartículas Metálicas/química , Microscopia Eletrônica de Transmissão e Varredura/métodos
18.
Macromol Rapid Commun ; 40(2): e1800333, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30027594

RESUMO

This article presents the synthesis of poly(methacrylic acid)-b-poly(methyl methacrylate) diblock copolymer via polymerization-induced self-assembly in the presence of iron-oxide nanoparticles. Detailed phase diagrams with and without inorganic nanoparticles were constructed. Scanning transmission electron microscopy and energy dispersive X-ray photometry studies confirme the decoration of the polymeric nanoparticles with the iron-oxide nanoparticles. These hybrid nanoparticles were used to prepare porous thin film membranes by spin coating. Finally, the magneto-responsive properties of the membranes were assessed using water filtration tests in the presence and absence of a magnetic field.


Assuntos
Técnicas de Química Sintética/métodos , Nanopartículas de Magnetita/química , Nanopartículas/química , Polímeros/química , Ácidos Polimetacrílicos/química , Polimetil Metacrilato/química , Materiais Revestidos Biocompatíveis/química , Campos Magnéticos , Nanopartículas de Magnetita/ultraestrutura , Membranas Artificiais , Microscopia Eletrônica de Transmissão e Varredura , Nanopartículas/ultraestrutura , Polimerização , Polímeros/síntese química , Porosidade
19.
Macromol Rapid Commun ; 40(2): e1800346, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29974542

RESUMO

The use of 2,3,4,5,6-pentafluorobenzyl methacrylate (PFBMA) as a core-forming monomer in ethanolic reversible addition-fragmentation chain transfer dispersion polymerization formulations is presented. Poly[poly(ethylene glycol) methyl ether methacrylate] (pPEGMA) macromolecular chain transfer agents were chain-extended with PFBMA leading to nanoparticle formation via polymerization-induced self-assembly (PISA). pPEGMA-pPFBMA particles exhibited the full range of morphologies (spheres, worms, and vesicles), including pure and mixed phases. Worm phases formed gels that underwent a thermo-reversible degelation and morphological transition to spheres (or spheres and vesicles) upon heating. Postsynthesis, the pPFBMA cores were modified through thiol-para-fluoro substitution reactions in ethanol using 1,8-diazabicyclo[5.4.0]undec-7-ene as the base. For monothiols, conversions were 64% (1-octanethiol) and 94% (benzyl mercaptan). Spherical and worm-shaped nano-objects were core cross-linked using 1,8-octanedithiol, which prevented their dissociation in nonselective solvents. For a temperature-responsive worm sample, cross-linking additionally resulted in the loss of the temperature-triggered morphological transition. The use of the reactive monomer PFBMA in PISA formulations presents a simple method to prepare well-defined nano-objects similar to those produced with nonreactive monomers (e.g., benzyl methacrylate) and to retain morphologies independent of solvent and temperature.


Assuntos
Álcoois Benzílicos/química , Técnicas de Química Sintética/métodos , Metacrilatos/química , Nanopartículas/química , Polimerização , Polímeros/química , Compostos de Sulfidrila/química , Compostos Bicíclicos Heterocíclicos com Pontes/química , Microscopia Eletrônica de Transmissão e Varredura , Modelos Químicos , Estrutura Molecular , Nanopartículas/ultraestrutura , Polímeros/síntese química , Temperatura , Temperatura de Transição
20.
Microscopy (Oxf) ; 68(2): 111-121, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30380073

RESUMO

A 3, 3'-diaminobenzidine (DAB)-based method was used to detect the localization of endogenous peroxidase activity in Indian rhinoceros (Rhinoceros unicornis) parotid gland acinar cells. The tissue had previously been resin-embedded in gelatin capsules for routine electron microscopic observations and thus pre-incubation for endogenous peroxidase analysis was not possible. We attempted to demonstrate the relationship between secretory granules (SGs) in resin ultrathin sections of Indian rhinoceros parotid gland acinar cells and endogenous peroxidase activity. A JEM 1400 Plus scanning transmission electron microscope (STEM) was used to conduct energy dispersive X-ray spectroscopy (EDS) analysis of the presence of nitrogen generated by the DAB reaction in bipartite structural SG consisting of a dense body (or core). The mapping patterns of nitrogen were restricted to the dense body. We observed nitrogen localized in the rough endoplasmic reticulum (ER), nuclear envelope (NE) and several components of the Golgi apparatus (G) of rhinoceros parotid gland acinar cells participating in the synthetic pathway of secretory proteins. Moreover, we established a nitrogen-detection method by EDS analysis of rhinoceros parotid gland. The reliability of the method was validated by comparison of the test group (peroxidase detection in ultrathin resin sections) and the control group (ordinary peroxidase detection in semi-thin sections following glutaraldehyde pre-fixation) of rat submandibular gland. The same mapping patterns of nitrogen were detected by DAB reaction in the SG, ER, NE and G in these two groups. Hence, EDS-STEM approaches for endogenous peroxidase post-incubation analysis will prove useful for advanced cytochemical analysis for the identification of any other resin sections.


Assuntos
3,3'-Diaminobenzidina/química , Células Acinares/ultraestrutura , Microscopia Eletrônica de Transmissão e Varredura/métodos , Nitrogênio/análise , Glândula Parótida/diagnóstico por imagem , Peroxidase/metabolismo , Animais , Retículo Endoplasmático/metabolismo , Feminino , Complexo de Golgi/metabolismo , Masculino , Microtomia/métodos , Membrana Nuclear/metabolismo , Glândula Parótida/citologia , Perissodáctilos , Ratos , Ratos Sprague-Dawley , Resinas Sintéticas/química , Fixação de Tecidos/métodos
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