scholarly journals Ionic Nanocomplexes of Hyaluronic Acid and Polyarginine to Form Solid Materials: A Green Methodology to Obtain Sponges with Biomedical Potential

Nanomaterials ◽  
2019 ◽  
Vol 9 (7) ◽  
pp. 944
Author(s):  
María Gabriela Villamizar-Sarmiento ◽  
Ignacio Moreno-Villoslada ◽  
Samuel Martínez ◽  
Annesi Giacaman ◽  
Victor Miranda ◽  
...  
Keyword(s):  
Hyaluronic Acid ◽  
High Capacity ◽  
Solid Material ◽  
Fibroblast Cell ◽  
High Porosity ◽  
Design Development ◽  
Solid Materials ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission

We report on the design, development, characterization, and a preliminary cellular evaluation of a novel solid material. This material is composed of low-molecular-weight hyaluronic acid (LMWHA) and polyarginine (PArg), which generate aqueous ionic nanocomplexes (INC) that are then freeze-dried to create the final product. Different ratios of LMWHA/PArg were selected to elaborate INC, the size and zeta potential of which ranged from 100 to 200 nm and +25 to −43 mV, respectively. Turbidimetry and nanoparticle concentration analyses demonstrated the high capacity of the INC to interact with increasing concentrations of LMWHA, improving the yield of production of the nanostructures. Interestingly, once the selected formulations of INC were freeze-dried, only those comprising a larger excess of LMWHA could form reproducible sponge formulations, as seen with the naked eye. This optical behavior was consistent with the scanning transmission electron microscopy (STEM) images, which showed a tendency of the particles to agglomerate when an excess of LMWHA was present. Mechanical characterization evidenced low stiffness in the materials, attributed to the low density and high porosity. A preliminary cellular evaluation in a fibroblast cell line (RMF-EG) evidenced the concentration range where swollen formulations did not affect cell proliferation (93–464 µM) at 24, 48, or 72 h. Considering that the reproducible sponge formulations were elaborated following inexpensive and non-contaminant methods and comprised bioactive components, we postulate them with potential for biomedical purposes. Additionally, this systematic study provides important information to design reproducible porous solid materials using ionic nanocomplexes.

Contrast and Resolution in Alfresco Microscopy and Thick Specimens

1972 ◽  
Vol 30 ◽  
pp. 570-571
Author(s):  
J. R. Sellar ◽  
J. M. Cowley
Keyword(s):  
Solid Material ◽  
Cell Design ◽  
Current Interest ◽  
High Voltage Electron Microscopy ◽  
Voltage Electron ◽  
Transmission Electron ◽  
Environmental Cell ◽  
Scanning Transmission ◽  
Conventional Transmission Electron Microscope ◽  
Transmission Microscope

Current interest in high voltage electron microscopy, especially in the scanning mode, has prompted the development of a method for determining the contrast and resolution of images of specimens in controlled-atmosphere stages or open to the air, hydrated biological specimens being a good example. Such a method would be of use in the prediction of microscope performance and in the subsequent optimization of environmental cell design for given circumstances of accelerating voltage, cell gas pressure and constitution, and desired resolution.Fig. 1 depicts the alfresco cell of a focussed scanning transmission microscope with a layer of gas L (and possibly a thin window W) between the objective O and specimen T. Using the principle of reciprocity, it may be considered optically equivalent to a conventional transmission electron microscope, if the beams were reversed. The layer of gas or solid material after the specimen in the STEM or before the specimen in TEM has no great effect on resolution or contrast and so is ignored here.

scholarly journals Development of nanostructured bioplastic material for wound healing

2021 ◽  
Author(s):  
Ilmira R. Gilmutdinova ◽  
Elena Kostromina ◽  
Regina D. Yakupova ◽  
Petr S. Eremin
Keyword(s):  
Hyaluronic Acid ◽  
Clinical Practice ◽  
Physical Properties ◽  
Human Fibroblasts ◽  
Fibroblast Cell ◽  
Human Skin Fibroblast ◽  
High Porosity ◽  
The Matrix ◽  
Photochemical Crosslinking ◽  
Fibroblast Cell Culture

The development of new biomaterials whose characteristics are as close as possible to the properties of living human tissues is one of the most promising areas of regenerative medicine. This work aimed at creating a bioplastic material based on collagen, elastin and hyaluronic acid and studying its structure and properties to assess the prospects for further use in clinical practice. Bioplastic material was obtained by mixing collagen, hyaluronic acid and elastin in predetermined proportions with distilled water. We treated the material with photochemical crosslinking to stabilize biofilm in a liquid medium and form a nanostructured scaffold. A commercial human skin fibroblast cell culture was used to assess the biomaterial cytotoxicity and biocompatibility. The visualization and studies of the biomaterial structure were performed using light and scanning electron microscopy. It has been shown that the obtained biomaterial is characterized by high resilience; it has also a high porosity. The co-culturing of the bioplastic material and human fibroblasts did not reveal any of its cytotoxic effects on cells in culture. It was shown that the biomaterial samples could maintain physical properties in the culture medium for more than 10 days, while the destruction of the matrix was observed 3–4 weeks after the beginning of incubation. Thus, the created biomaterial can be used on damaged skin areas due to its physical properties and structure. The use of the developed biomaterial provides effective conditions for good cell proliferation, which allows us to consider it as a promising wound cover for use in clinical practice.

scholarly journals Development of nanostructured bioplastic material for wound healing

2021 ◽  
Author(s):  
Ilmira R. Gilmutdinova ◽  
Elena Kostromina ◽  
Regina D. Yakupova ◽  
Petr S. Eremin
Keyword(s):  
Hyaluronic Acid ◽  
Clinical Practice ◽  
Physical Properties ◽  
Human Fibroblasts ◽  
Fibroblast Cell ◽  
Human Skin Fibroblast ◽  
High Porosity ◽  
The Matrix ◽  
Photochemical Crosslinking ◽  
Fibroblast Cell Culture

The development of new biomaterials whose characteristics are as close as possible to the properties of living human tissues is one of the most promising areas of regenerative medicine. This work aimed at creating a bioplastic material based on collagen, elastin and hyaluronic acid and studying its structure and properties to assess the prospects for further use in clinical practice. Bioplastic material was obtained by mixing collagen, hyaluronic acid and elastin in predetermined proportions with distilled water. We treated the material with photochemical crosslinking to stabilize biofilm in a liquid medium and form a nanostructured scaffold. A commercial human skin fibroblast cell culture was used to assess the biomaterial cytotoxicity and biocompatibility. The visualization and studies of the biomaterial structure were performed using light and scanning electron microscopy. It has been shown that the obtained biomaterial is characterized by high resilience; it has also a high porosity. The co-culturing of the bioplastic material and human fibroblasts did not reveal any of its cytotoxic effects on cells in culture. It was shown that the biomaterial samples could maintain physical properties in the culture medium for more than 10 days, while the destruction of the matrix was observed 3–4 weeks after the beginning of incubation. Thus, the created biomaterial can be used on damaged skin areas due to its physical properties and structure. The use of the developed biomaterial provides effective conditions for good cell proliferation, which allows us to consider it as a promising wound cover for use in clinical practice.

Structural analysis of biological macromolecular assemblies by scanning transmission electron microscopy

1991 ◽  
Vol 49 ◽  
pp. 262-263
Author(s):  
M. Boublik ◽  
S. J. Tumminia ◽  
W. Hellmann ◽  
Q. Zhang ◽  
J.F. Hainfeld ◽  
...  
Keyword(s):  
Signal To Noise Ratio ◽  
Macromolecular Complex ◽  
Functional Sites ◽  
Macromolecular Assemblies ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
Annular Detector ◽  
Topographical Mapping ◽  
Particle Elimination

Separation of the resolution and contrast affecting components and the optimized placement of detectors for the collection of elastic (contrast-forming) electrons on Brookhaven dedicated STEM make it possible to quantitatively detect greater than 90% of the available elastically scattered electrons. The high contrast and superior signal-to-noise ratio associated with the STEM annular detector allow for the imaging of unstained freeze-dried biological macromolecular complexes (chromatin, viruses, nucleic acids) at radiation doses as low as 1 e/Å. Specimens prepared in this way are free of the main resolution-limiting conditions of conventional TEM i.e. staining , air drying and radiation damage. The image intensity of unstained specimens can be related to their local projected mass and used for calculation of the total mass and mass distribution within any selected particle. Elimination of staining makes it possible to use heavy metals as high-resolution markers for topographical mapping of components and/or functional sites on a particular macromolecular complex.

Higher-order assembly of gastropodan hemocyanins as revealed by Scanning Transmission Electron Microscopy (STEM)

1989 ◽  
Vol 47 ◽  
pp. 248-249
Author(s):  
M.G. Hamilton ◽  
R.R. Rodriguez ◽  
T.T. Herskovits ◽  
J.S. Wall
Keyword(s):  
Analytical Ultracentrifugation ◽  
Large Angle ◽  
Gel Filtration ◽  
Scattered Electron ◽  
Molecular Weights ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
The Masses ◽  
Individual Particles

The hemocyanins of gastropods consist of aggregates of a cylindrical decameric subparticle that assembles into di-, tri-, tetra-, penta-, and larger multi-decameric particles with sedimentation coefficients of ca. 105 S, 130 S, 150 S, 170 S, and higher values. We are using STEM to measure the masses of individual particles and analytical ultracentrifugation to determine the distribution of sedimenting components.Hemocyanins were isolated from freshly collected hemolymph by gel filtration on BioGel A-5m columns. Samples were analyzed with schlieren optics in a Beckman Madel E ultracentrifuge. Specimens were diluted into 0.1 M HEPES, pH 8.0, 0.01 M MgC12 to a final concentration of 100 ug/mL and freeze-dried for STEM analysis. The STEM instrument was operated at 40 kV using a -140 °C cold stage. The elastically scattered electron signal from the STEM large angle annular detector was used to form the images. The specimens were imaged with 10 A pixels at a dose of 6-10 e/A2. Molecular weights of individual particles were measured as previously described.

scholarly journals Solution and surface effects on plasma fibronectin structure.

1983 ◽  
Vol 97 (6) ◽  
pp. 1686-1692 ◽  
Author(s):  
N M Tooney ◽  
M W Mosesson ◽  
D L Amrani ◽  
J F Hainfeld ◽  
J S Wall
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Ionic Strength ◽  
Scanning Transmission Electron Microscopy ◽  
Plasma Fibronectin ◽  
Scanning Transmission Electron ◽  
Neutral Ph ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission

As assessed by electron microscopy, the reported shape of the plasma fibronectin molecule ranges from that of a compact particle to an elongated, rod-like structure. In this study, we evaluated the effects of solution and surface conditions on fibronectin shape. Freeze-dried, unstained human plasma fibronectin molecules deposited at pH 7.0-7.4 onto carbon films and examined by scanning transmission electron microscopy appeared relatively compact and pleiomorphic, with approximate average dimensions of 24 nm X 16 nm. Negatively stained molecules also had a similar shape but revealed greater detail in that we observed irregular, yarn-like structures. Glutaraldehyde-induced intramolecular cross-linking did not alter the appearance of plasma fibronectin. Molecules deposited at pH 2.8, pH 9.3, or after succinylation were less compact than those deposited at neutral pH. In contrast, fibronectin molecules sprayed onto mica surfaces at pH 7, rotary shadowed, and examined by transmission electron microscopy were elongated and nodular with a contour length of 120-130 nm. Sedimentation velocity experiments and electron microscopic observations indicate that fibronectin unfolds when it is succinylated, when the ionic strength is raised at pH 7, or when the pH is adjusted to 9.3 or 2.8. Greater unfolding is observed at pH 2.8 at low ionic strength (less than 0.01) compared with material at that pH in 0.15 M NaCl solution. We conclude that (a) the shape assumed by the fibronectin molecule can be strongly affected by solution conditions and by deposition onto certain surfaces; and that (b) the images of fibronectin seen by scanning transmission electron microscopy at neutral pH on carbon film are representative of molecules in physiologic solution.

X-ray microanalysis of mineralized deposits in a variety of pathological conditions in the brain

1995 ◽  
Vol 53 ◽  
pp. 866-867
Author(s):  
C. A. Ackerley ◽  
L. E. Becker
Keyword(s):  
Electron Microscope ◽  
Scanning Transmission Electron Microscope ◽  
Small Degree ◽  
X Ray ◽  
Transmission Electron ◽  
Formalin Fixed Paraffin ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
Formalin Fixed Paraffin Embedded ◽  
The Brain

Although a small degree of mineralization can be a common occurrence without associated pathological symptoms, certain diseases of the brain do however exhibit distinct increases in mineralization with characteristic distributions l>2. In this study, tissues from a number of these disorders were prepared for x-ray microanalysis in several ways. Where possible, material was slam frozen on a liquid nitrogen cooled polished copper block, cryosections prepared and freeze dried in the scanning transmission electron microscope (STEM) using a cold stage prior to analysis by energy dispersive x-ray spectrometry (EDS). In addition, samples were freeze substituted for several days, embedded in LR white and cut on dry knives before analysis. Where only formalin fixed paraffin embedded materials were available, .5μ.m sections were cut and mounted on carbon planchets. The specimens were then deparaffinized with xylene and viewed with the backscatter electron detector (BEI) in the scanning electron microscope (SEM) and analyzed by EDS.

Ion localization in Cells by Electron Probe X-Ray Microanalysis (Epxma) Depends on Cryopreparation

1990 ◽  
Vol 48 (2) ◽  
pp. 322-323
Author(s):  
Karl Zierold
Keyword(s):  
Detection Limit ◽  
Scanning Transmission Electron Microscopy ◽  
Dry Weight ◽  
Section Thickness ◽  
X Ray ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
Absolute Detection Limit ◽  
Analytical Resolution

Elements in biological cells can be localized by EPXMA. The most attractive approach of this method with respect to detection limit and spatial resolution is scanning transmission electron microscopy combined with energy dispersive x ray microanalysis of ultrathin (approximately 100 nm thick) freeze-dried cryosections. The detection limit, here determined by scanning an electron beam of 1.3 nA for 2 min over freeze-dried cryosections from frozen standard solutions was found to be about 10 mMol/kg dry weight for all elements with the atomic number Z higher than 12. Due to the Be window in the x ray detector the detection limit was 20 mMol/kg dry weight for Mg (Z = 12) and 30 mMol/kg dry weight for Na (Z = 11). The lateral analytical resolution was less than 50 nm, limited by section thickness. In most cells 10 mMol/kg dry weight correspond to an absolute detection limit of .500 atoms within a volume of 100×50×50 nm3.

Structural Analysis of Filaments Using the STEM

2000 ◽  
Vol 6 (S2) ◽  
pp. 862-863
Author(s):  
M. N. Simon ◽  
B. Y. Lin ◽  
J. S. Wall
Keyword(s):  
Scanning Transmission Electron Microscope ◽  
Digital Data ◽  
Mass Analysis ◽  
Biological Origin ◽  
Biotechnology Research ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Different Types ◽  
Scanning Transmission ◽  
History Of

The Scanning Transmission Electron Microscope (STEM) facility at BNL is an NIH Biotechnology Research Resource and as such is available to users with suitable projects, free of charge. Currently, many of our users' projects involve studying the structures of a variety of filaments. Most of these are of biological origin, although a couple involve conducting polymers. The STEM has a long history of being used to study different types of filaments and resolving controversies about their structure.The mainstay of the STEM is mass analysis on unstained, isolated, freeze-dried samples. On these, the STEM can collect in-focus digital data directly. In a scan (8 sec. in real time) of a sample, at each of 512x512 picture elements (pixels), the number of electrons scattered into two annular detectors is recorded. For each pixel, the number of scattered electrons is directly proportional to the mass thickness in that pixel.

Analysis of Molluscan Hemocyanins by Scanning Transmission Electron Microscopy and Light Scattering

1988 ◽  
Vol 46 ◽  
pp. 162-163
Author(s):  
P. S. Furcinitti ◽  
J. S. Wall ◽  
M. G. Hamilton ◽  
T. T. Herskovits
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Light Scattering ◽  
Scanning Transmission Electron Microscopy ◽  
Large Angle ◽  
Scattered Electron ◽  
Scanning Transmission Electron ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission

Hemocyanins are copper containing, oxygen binding proteins found in many invertebrate species of the phyla Arthropoda and Mollusca. Molluscan hemocyanins are cylindrical macromolecular assemblies of a basic decameric unit. The hemocyanins of two chitons, Stenonlax conspicua and Mopalia muscosa. are decamers, while those of two gastropods, Fasciolaria tulipa and Pleuroplora gieantea. are di-decamers. The hemocyanins of a third gastropod species, Busvcon contrarium can exist in a spectrum of multi-decameric forms. Molecular weights of the various types of molluscan hemocyanins were measured by absolute light scattering and Scanning Transmission Electron Microscopy (STEM) as a first step in understanding multi-decamer formation in some types of hemocyanins.Hemocyanins were prepared as previously described, dialyzed into 1M HEPES buffer, pH 7.4, containing 0.05M MgCl and freeze-dried or negatively stained for STEM analysis. Specimens were examined at the Brookhaven STEM Biotechnology Resource, which was operated at 40 kV using a -140°C cold stage. The elastically scattered electron signal from the STEM large angle annular detector was used to form the images.

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