scholarly journals Effect of enzymic (collagenase) harvesting on the intracellular Na+/K+ ratio of Swiss/3T3 cells as revealed by X-ray microanalysis

1988 ◽  
Vol 90 (1) ◽  
pp. 99-104
Author(s):  
Z. Szallasi ◽  
A. Szallasi ◽  
F. Bojan ◽  
I. Zs-Nagy
Keyword(s):  
Cell Cultures ◽  
Freeze Fracture ◽  
Ion Concentration ◽  
Bulk Specimen ◽  
X Ray ◽  
Dependent Cell ◽  
Swiss 3T3 ◽  
K Concentration ◽  
Post Harvesting ◽  
Swiss 3T3 Cell

Swiss/3T3 cell cultures were harvested with 0.05% collagenase and after centrifugation the pellet was prepared by the freeze-fracture/freeze-drying (FFFD) method for bulk-specimen X-ray microanalysis. Time-dependent variations in the intracellular monovalent elemental concentrations (Na+, K+ and Cl-) as well as of the Na+/K+ ratio were followed for 120 min subsequent to harvesting. The quantitative measurements revealed a very considerable increase in the intracellular Na+ and Cl- accompanied by a decrease in the K+ concentration as soon as 5 min after harvesting. The Na+/K+ ratio had increased by this time to about 1.5 on average. These changes indicate a sustained depolarization of the cell membrane. During the first 60 min this depolarization tended to normalize as demonstrated by an exponential decrease in the intracellular Na+ and Cl- and an increase in the K+ content involving a decrease in the Na+/K+ ratio. The total intracellular monovalent ion concentration remained almost constant during this post-harvesting period. These results suggest that harvesting represents a serious depolarizing stimulus to the cells, the consequences of which are restored only after 1–2h. These alterations should be taken into consideration during various experimental designs when using anchorage-dependent cell cultures.

Direct visualization of phase-separated domains in lipid membranes

1978 ◽  
Vol 36 (2) ◽  
pp. 290-291
Author(s):  
S. W. Hui ◽  
T. P. Stewart
Keyword(s):  
Lipid Membranes ◽  
Structural Information ◽  
Freeze Fracture ◽  
Biological Molecules ◽  
Vacuum Drying ◽  
Direct Visualization ◽  
X Ray Diffraction ◽  
Electron Microscopic ◽  
X Ray ◽  
Hydrocarbon Chains

Direct electron microscopic study of biological molecules has been hampered by such factors as radiation damage, lack of contrast and vacuum drying. In certain cases, however, the difficulties may be overcome by using redundent structural information from repeating units and by various specimen preservation methods. With bilayers of phospholipids in which both the solid and fluid phases co-exist, the ordering of the hydrocarbon chains may be utilized to form diffraction contrast images. Domains of different molecular packings may be recgnizable by placing properly chosen filters in the diffraction plane. These domains would correspond to those observed by freeze fracture, if certain distinctive undulating patterns are associated with certain molecular packing, as suggested by X-ray diffraction studies. By using an environmental stage, we were able to directly observe these domains in bilayers of mixed phospholipids at various temperatures at which their phases change from misible to inmissible states.

Freeze-Fracture Replication of Frozen Outer Segments of Rat Retinal Rods

1969 ◽  
Vol 27 ◽  
pp. 392-393
Author(s):  
Thomas S. Leeson ◽  
C. Roland Leeson
Keyword(s):  
Electron Microscopy ◽  
Cross Section ◽  
Outer Segment ◽  
Freeze Fracture ◽  
X Ray Diffraction ◽  
X Ray ◽  
Outer Segments ◽  
Retinal Rods ◽  
Temperature Electron ◽  
Freeze Etching

Numerous previous studies of outer segments of retinal receptors have demonstrated a complex internal structure of a series of transversely orientated membranous lamellae, discs, or saccules. In cones, these lamellae probably are invaginations of the covering plasma membrane. In rods, however, they appear to be isolated and separate discs although some authors report interconnections and some continuities with the surface near the base of the outer segment, i.e. toward the inner segment. In some species, variations have been reported, such as longitudinally orientated lamellae and lamellar whorls. In cross section, the discs or saccules show one or more incisures. The saccules probably contain photolabile pigment, with resulting potentials after dipole formation during bleaching of pigment. Continuity between the lamina of rod saccules and extracellular space may be necessary for the detection of dipoles, although such continuity usually is not found by electron microscopy. Particles on the membranes have been found by low angle X-ray diffraction, by low temperature electron microscopy and by freeze-etching techniques.

Techniques for cryoultramicrotomy of propane jet frozen biological samples

1986 ◽  
Vol 44 ◽  
pp. 260-261
Author(s):  
B. Craig ◽  
L. Hawkey ◽  
A. LeFurgey
Keyword(s):  
Freeze Fracture ◽  
Freeze Substitution ◽  
Heart Cells ◽  
Crystal Formation ◽  
X Ray ◽  
Transmission Electron ◽  
Scanning Transmission ◽  
Chick Heart ◽  
Embryonic Chick Heart ◽  
A New Technique

Ultra-rapid freezing followed by cryoultramicrotomy is essential for the preservation of diffusible elements in situ within cells prior to scanning transmission electron microscopy and quantitative energy dispersive x-ray microanalysis. For cells or tissue fragments in suspension and for monolayer cell cultures, propane jet freezing provides cooling rates greater than 30,000°C/sec with regions up to 40μm in thickness free of significant ice crystal formation. While this method of freezing has frequently been applied prior to freeze fracture or freeze substitution, it has not been widely utilized prior to cryoultramicrotomy and subsequent x-ray microanalytical studies. This report describes methods devised in our laboratory for cryosectioning of propane jet frozen kidney proximal tubule suspensions and cultured embryonic chick heart cells, in particular a new technique for mounting frozen suspension specimens for sectioning. The techniques utilize the same specimen supports and sample holders as those used for freeze fracture and freeze substitution and should be generally applicable to any cell suspension or culture preparation.

scholarly journals DNA Aptamer Functionalized Hydrogels for Interferometric Fiber-Optic Based Continuous Monitoring of Potassium Ions

Biosensors ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 266
Author(s):  
Nataša Žuržul ◽  
Bjørn Torger Stokke
Keyword(s):  
Conformational Transition ◽  
Polymer Network ◽  
Potassium Ion ◽  
Ion Concentration ◽  
Dna Aptamer ◽  
Blood Levels ◽  
Label Free ◽  
Practical Applications ◽  
Hydrogel Matrix ◽  
K Concentration

In the present paper, we describe a potassium sensor based on DNA-aptamer functionalized hydrogel, that is capable of continuous label-free potassium ion (K+) monitoring with potential for in situ application. A hydrogel attached to the end of an optical fiber is designed with di-oligonucleotides grafted to the polymer network that may serve as network junctions in addition to the covalent crosslinks. Specific affinity toward K+ is based on exploiting a particular aptamer that exhibits conformational transition from single-stranded DNA to G-quadruplex formed by the di-oligonucleotide in the presence of K+. Integration of this aptamer into the hydrogel transforms the K+ specific conformational transition to a K+ concentration dependent deswelling of the hydrogel. High-resolution interferometry monitors changes in extent of swelling at 1 Hz and 2 nm resolution for the hydrogel matrix of 50 µm. The developed hydrogel-based biosensor displayed high selectivity for K+ ions in the concentration range up to 10 mM, in the presence of physiological concentrations of Na+. Additionally, the concentration dependent and selective K+ detection demonstrated in the artificial blood buffer environment, both at room and physiological temperatures, suggests substantial potential for practical applications such as monitoring of potassium ion concentration in blood levels in intensive care medicine.

scholarly journals Pristine and Magnetic Kenaf Fiber Biochar for Cd2+ Adsorption from Aqueous Solution

2021 ◽  
Vol 18 (15) ◽  
pp. 7949
Author(s):  
Anwar Ameen Hezam Saeed ◽  
Noorfidza Yub Harun ◽  
Suriati Sufian ◽  
Muhammad Roil Bilad ◽  
Zaki Yamani Zakaria ◽  
...  
Keyword(s):  
Adsorption Capacity ◽  
Photoelectron Spectroscopy ◽  
Textural Properties ◽  
Ion Concentration ◽  
Solution Temperature ◽  
Maximum Adsorption Capacity ◽  
Magnetic Biochar ◽  
X Ray ◽  
Study Results ◽  
Capacity Surface

Development of strategies for removing heavy metals from aquatic environments is in high demand. Cadmium is one of the most dangerous metals in the environment, even under extremely low quantities. In this study, kenaf and magnetic biochar composite were prepared for the adsorption of Cd2+. The synthesized biochar was characterized using (a vibrating-sample magnetometer VSM), Scanning electron microscopy (SEM), X-ray powder diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), and X-ray photoelectron spectroscopy (XPS). The adsorption batch study was carried out to investigate the influence of pH, kinetics, isotherm, and thermodynamics on Cd2+ adsorption. The characterization results demonstrated that the biochar contained iron particles that help in improving the textural properties (i.e., surface area and pore volume), increasing the number of oxygen-containing groups, and forming inner-sphere complexes with oxygen-containing groups. The adsorption study results show that optimum adsorption was achieved under pH 5–6. An increase in initial ion concentration and solution temperature resulted in increased adsorption capacity. Surface modification of biochar using iron oxide for imposing magnetic property allowed for easy separation by external magnet and regeneration. The magnetic biochar composite also showed a higher affinity to Cd2+ than the pristine biochar. The adsorption data fit well with the pseudo-second-order and the Langmuir isotherm, with the maximum adsorption capacity of 47.90 mg/g.

scholarly journals Magnetically Recyclable Wool Keratin Modified Magnetite Powders for Efficient Removal of Cu2+ Ions from Aqueous Solutions

Nanomaterials ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1068
Author(s):  
Xinyue Zhang ◽  
Yani Guo ◽  
Wenjun Li ◽  
Jinyuan Zhang ◽  
Hailiang Wu ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Aqueous Solutions ◽  
Adsorption Capacity ◽  
Surface Area ◽  
Ion Concentration ◽  
Bet Surface Area ◽  
Chemical Compositions ◽  
Ion Adsorption ◽  
X Ray ◽  
Wool Keratin

The treatment of wastewater containing heavy metals and the utilization of wool waste are very important for the sustainable development of textile mills. In this study, the wool keratin modified magnetite (Fe3O4) powders were fabricated by using wool waste via a co-precipitation technique for removal of Cu2+ ions from aqueous solutions. The morphology, chemical compositions, crystal structure, microstructure, magnetism properties, organic content, and specific surface area of as-fabricated powders were systematically characterized by various techniques including field emission scanning electron microscopy (FESEM), energy dispersive spectroscopy (EDS), X-ray diffraction (XRD), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), vibrating sample magnetometer (VSM), thermogravimetric (TG) analysis, and Brunauer–Emmett–Teller (BET) surface area analyzer. The effects of experimental parameters such as the volume of wool keratin hydrolysate, the dosage of powder, the initial Cu2+ ion concentration, and the pH value of solution on the adsorption capacity of Cu2+ ions by the powders were examined. The experimental results indicated that the Cu2+ ion adsorption performance of the wool keratin modified Fe3O4 powders exhibited much better than that of the chitosan modified ones with a maximum Cu2+ adsorption capacity of 27.4 mg/g under favorable conditions (0.05 g powders; 50 mL of 40 mg/L CuSO4; pH 5; temperature 293 K). The high adsorption capacity towards Cu2+ ions on the wool keratin modified Fe3O4 powders was primarily because of the strong surface complexation of –COOH and –NH2 functional groups of wool keratins with Cu2+ ions. The Cu2+ ion adsorption process on the wool keratin modified Fe3O4 powders followed the Temkin adsorption isotherm model and the intraparticle diffusion and pseudo-second-order adsorption kinetic models. After Cu2+ ion removal, the wool keratin modified Fe3O4 powders were easily separated using a magnet from aqueous solution and efficiently regenerated using 0.5 M ethylene diamine tetraacetic acid (EDTA)-H2SO4 eluting. The wool keratin modified Fe3O4 powders possessed good regenerative performance after five cycles. This study provided a feasible way to utilize waste wool textiles for preparing magnetic biomass-based adsorbents for the removal of heavy metal ions from aqueous solutions.

Thin-section, freeze-fracture, and energy dispersive x-ray analysis studies of the protein bodies of tomato seeds

1980 ◽  
Vol 58 (6) ◽  
pp. 699-711 ◽  
Author(s):  
Ernest Spitzer ◽  
John N. A. Lott
Keyword(s):  
Freeze Fracture ◽  
Protein Body ◽  
Cell Types ◽  
Energy Dispersive ◽  
Protein Bodies ◽  
Elemental Content ◽  
Tomato Seed ◽  
Endosperm Tissue ◽  
X Ray ◽  
Edx Analysis

Protein bodies of dry seeds of tomato (Lycopersicon esculentum) from radicle, hypocotyl, cotyledon, and endosperm tissue were extensively studied using thin-sectioning, freeze-fracturing and energy dispersive x-ray (EDX) analysis. Protein bodies varied in size, were oval to circular in section, and generally consisted of a proteinaceous matrix, globoid crystal, and protein crystalloid components. Size, shape, and arrangements of globoid crystals and protein crystalloids varied even within the same cell. Globoid crystals were generally oval to circular in section. They were always surrounded by a proteinaceous matrix. In a given protein body the number present ranged from a few to numerous. A protein body generally contained only one protein crystalloid. In section, protein crystalloids were irregular or angular in shape. They were composed of substructural particles which formed lattice planes. EDX analysis of tomato seed globoid crystals revealed the presence of P, K, and Mg in all cases, a fact that is consistent with globoid crystals being phytin-rich. Rarely, small amounts of calcium were found along with P, K, and Mg in globoid crystals of each of the tissue regions considered. The distribution pattern of cells with Ca containing globoid crystals was random. Small amounts of Fe and Mn were also found in the globoid crystals of protein bodies from certain cell types. These two elements, unlike calcium, were specific in terms of their distribution. Globoid crystals from the protodermal cells often contained Mn and Fe. The globoid crystals from provascular tissue of radicle, hypocotyl, and cotyledon regions often contained Fe while globoid crystals in the first layer of large cells surrounding these provascular areas always contained Fe. Results from EDX analysis of the proteinaceous material from the protein bodies are presented and discussed as are variations in elemental content due to different fixations.

An improved approach to freeze-fracture morphology of monolayer cell cultures

1998 ◽  
Vol 82 (1) ◽  
pp. 97-103 ◽  
Author(s):  
Elisabeth Bugnard ◽  
Patrick Sors ◽  
Alain Bloc ◽  
Françoise Loctin ◽  
Yves Dunant
Keyword(s):  
Cell Cultures ◽  
Freeze Fracture ◽  
Fracture Morphology ◽  
Monolayer Cell

scholarly journals Osmotic shrinkage of sterically stabilized liposomes as revealed by time-resolved small-angle X-ray scattering

2014 ◽  
Vol 47 (1) ◽  
pp. 35-40 ◽  
Author(s):  
Zoltán Varga ◽  
András Wacha ◽  
Attila Bóta
Keyword(s):  
Form Factor ◽  
Small Angle ◽  
Freeze Fracture ◽  
Initial Step ◽  
Time Resolved ◽  
X Ray ◽  
X Ray Scattering ◽  
Sterically Stabilized Liposomes ◽  
Liposome Size ◽  
Ray Scattering

Time-resolved synchrotron small-angle X-ray scattering (SAXS) was used to study the structural changes during the osmotic shrinkage of a pharmacologically relevant liposomal drug delivery system. Sterically stabilized liposomes (SSLs) with a diameter of 100 nm and composed of hydrogenated soy phosphocholine, cholesterol and distearoyl-phosphoethanolamine-PEG 2000 prepared in a salt-free buffer were mixed with a buffered 0.3 MNaCl solution using a stopped flow apparatus. The changes in the liposome size and the bilayer structure were followed by using SAXS with a time resolution of 20 ms. A linear decrease in liposome size is observed during the first ∼4 s of the osmotic shrinkage, which reveals a water permeability value of 0.215 (15) µm s−1. The change in the size of the liposomes upon the osmotic shrinkage is also confirmed by dynamic light scattering. After this initial step, broad correlation peaks appear on the SAXS curves in theqrange of the bilayer form factor, which indicates the formation of bi- or oligolamellar structures. Freeze-fracture combined with transmission electron microscopy revealed that lens-shaped liposomes are formed during the shrinkage, which account for the appearance of the quasi-Bragg peaks superimposed on the bilayer form factor. On the basis of these observations, it is proposed that the osmotic shrinkage of SSLs is a two-step process: in the initial step, the liposome shrinks in size, while the area/lipid adapts to the decreased surface area, which is then followed by the deformation of the spherical liposomes into lens-shaped vesicles.

scholarly journals Cloning, expression, purification, crystallization and preliminary X-ray diffraction of a lysine-specific permease fromPseudomonas aeruginosa

2014 ◽  
Vol 70 (10) ◽  
pp. 1362-1367 ◽  
Author(s):  
Emmanuel Nji ◽  
Dianfan Li ◽  
Declan A. Doyle ◽  
Martin Caffrey
Keyword(s):  
Metal Ion ◽  
Cell Volume Regulation ◽  
Ion Concentration ◽  
Size Exclusion ◽  
Substrate Selectivity ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Exclusion Chromatography

The prokaryotic lysine-specific permease (LysP) belongs to the amino acid–polyamine–organocation (APC) transporter superfamily. In the cell, members of this family are responsible for the uptake and recycling of nutrients, for the maintenance of a constant internal ion concentration and for cell volume regulation. The detailed mechanism of substrate selectivity and transport of L-lysine by LysP is not understood. A high-resolution crystal structure would enormously facilitate such an understanding. To this end, LysP fromPseudomonas aeruginosawas recombinantly expressed inEscherichia coliand purified to near homogeneity by immobilized metal ion-affinity chromatography (IMAC) and size-exclusion chromatography (SEC). Hexagonal- and rod-shaped crystals were obtained in the presence of L-lysine and the L-lysine analogue L-4-thialysine by vapour diffusion and diffracted to 7.5 Å resolution. The diffraction data were indexed in space groupP21, with unit-cell parametersa= 169.53,b= 169.53,c= 290.13 Å, γ = 120°.

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