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Fused-silica capillaries for capillary electrophoresis and gas chromatography: inner surface corrosion, within-batch differences, and influence of drawing parameters studied by atomic force microscopy.

Fused-silica capillaries for capillary electrophoresis (CE) and gas chromatography (GC) were investigated by atomic force microscopy (AFM). Differences from batch to batch and within one batch were often observed. Surface heterogeneity can be caused by bulk material, manufacturing parameters, or by aging effects. One batch of a fused-silica capillary was stored in water for three years at room temperature. The significant increase in surface roughness (measured as rms = root mean square) during this time is demonstrated. The effect of different drawing temperatures was investigated. Other drawing parameters were kept constant using one capillary batch. If the chosen drawing temperature was too low, the roughness values more than doubled. This increase in roughness did not affect the separation efficiency. However, the relative standard deviation (RSD%) of migration times and peak areas increased at the same time. Three capillary batches for gas chromatography of different inner diameters (250 microm, 320 microm, 530 microm) were also investigated. In all cases the higher rms values for surface roughness could be found at the beginning of the drawing process, although all values were close to atomic flatness.

Chromatography, Gas↗

Atomic force microscopy imaging of actin cortical cytoskeleton of Xenopus laevis oocyte.

In this study we report an atomic force microscopy (AFM) investigation of the actin cortical cytoskeleton of Xenopus laevis oocytes. Samples consisted of inside-out orientated plasma membrane patches of X. laevis oocytes with overhanging cytoplasmic material. They were spread on a freshly cleaved mica surface, subsequently treated with Triton X-100 detergent and chemically fixed. The presence of actin fibres in oocyte patches was proved by fluorescence microscopy imaging. Contact mode AFM imaging was performed in air in constant force conditions. Reproducible high-resolution AFM images of a filamentous structure were obtained. The filamentous structure was identified as an actin cortical cytoskeleton, investigating its disaggregation induced by cytochalasin D treatment. The thinnest fibres showed a height of 7 nm in accordance with the diameter of a single actin microfilament. The results suggest that AFM imaging can be used for the high-resolution study of the actin cortical cytoskeleton of the X. laevis oocyte and its modifications mediated by the action of drugs and toxins.

Actins↗

Atomic force microscopy studies of conditioner thickness distribution and binding interactions on the hair surface.

The way in which common hair care products, such as conditioner, deposit onto and change hair properties is of interest in beauty care science, as these properties are closely tied to product performance. The binding interaction between conditioner and the hair surface is one of the important factors in determining the conditioner thickness distribution and consequently the proper functions of conditioner. In this study, atomic force microscopy was used to obtain the local conditioner thickness distribution, adhesive forces and effective Young's modulus mapping of various hair surfaces. The conditioner thickness was extracted by measuring the forces on the atomic force microscopy tip as it approached, contacted and pushed through the conditioner layer. The effective Young's moduli of various hair surfaces were calculated from the force distance curves using Hertz analysis. The intrinsic binding interactions between different silicones and the hair surface on the microscopic scale, as well as their effect on the effective Young's modulus of the hair, are also discussed. It was found that the effective Young's modulus of the hair is strongly affected by the binding of conditioner molecules on the hair surface.

Hair↗

Atomic force microscopy can be used to mechanically stimulate osteoblasts and evaluate cellular strain distributions.

In this study, atomic force microscopy (AFM) was used to mechanically stimulate primary osteoblasts. In response to mechanical force applied by the AFM, the indented cell increased its intracellular calcium concentration. The material properties of the cell could be estimated and the membrane strains calculated. We proceeded to validate this technique experimentally and a 20% error was found between the predicted and the measured diameter of indentation. We also determined the strain distributions within the cell that result from AFM indentation using a simple finite element model. This enabled us to formulate hypotheses as to the mechanism through which cells may sense the applied mechanical strains. Finally, we report the effect of the Poisson ratio and the cell thickness on the strain distributions. Varying the Poisson ratio did not change the order of magnitude of the strains; whereas the cellular thickness dramatically changed the order of magnitude of the cellular strains. We conclude that AFM can be used for controlled mechanical stimulation of osteoblasts and that cellular strain distributions can be computed with a good accuracy when the cell is indented in its highest part.

Animals↗

Atomic force microscopy of paired helical filaments isolated from the autopsied brains of patients with Alzheimer's disease and immunolabeled against microtubule-associated protein tau.

Atomic force microscopy was employed to study the structural features of paired helical filaments isolated from autopsied brains of Alzheimer's disease patients. The identity of paired helical filaments was confirmed following a specific immunogold labeling using antibodies directed against the microtubule-associated protein tau, which is the main constituent of paired helical filaments. Computer-assisted analysis of high resolution, three-dimensional images allowed us to study the longitudinal and cross-sectional profiles of individual filaments. Vertical dimensions of filaments were assessed along these sectional profiles. The smallest vertical diameter (6.66 +/- 0.78 nm) was obtained at the level of the greatest lateral profile (ie, "loop"), while the greatest vertical diameter was two times larger (13.68 +/- 1.46 nm) and was obtained at the level of the smallest lateral profile (ie, "crossover") of the filament. Based on the shapes of these sectional profiles and their vertical dimensions, paired helical filaments appeared to be composed of two identical integral subunits, each of a circular cross-sectional profile of approximately 7 nm in diameter, wound around one another in a left helical manner, with a 7-nm center-to-center separation. Half-period of this helix was estimated at 81.4 +/- 2.1 nm. Serial cross-sectional profiles of paired helical filaments were further utilized to construct a theoretical model of their internal organization. This model suggests that each structural subunit of the paired helical filament incorporates at least four identical protofilaments.

Alzheimer Disease↗

Imaging saponin-induced structural changes in neural processes with atomic force microscopy.

Temporal changes in the structure of neuronal processes in the presence of saponin were studied by atomic force microscopy in a fluid medium. After saponin treatment, concavities were formed on the surface of some neurites and fibrous structures in other neurites were splayed. The vertical height of these splayed fibrils or fibrillar bundles ranged from 13 to 370 nm, and the horizontal width was less than 500 nm. These findings suggest that formation of concavities and separation of bundled fibrils occurred simultaneously in saponin-treated neurites.

Animals↗

Atomic force microscopy sees nucleosome positioning and histone H1-induced compaction in reconstituted chromatin.

We addressed the question of how nuclear histones and DNA interact and form a nucleosome structure by applying atomic force microscopy to an in vitro reconstituted chromatin system. The molecular images obtained by atomic force microscopy demonstrated that oligonucleosomes reconstituted with purified core histones and DNA yielded a 'beads on a string' structure with each nucleosome trapping 158 +/- 27 bp DNA. When dinucleosomes were assembled on a DNA fragment containing two tandem repeats of the positioning sequence of the Xenopus 5S RNA gene, two nucleosomes were located around each positioning sequence. The spacing of the nucleosomes fluctuated in the absence of salt and the nucleosomes were stabilized around the range of the positioning signals in the presence of 50 mM NaCl. An addition of histone H1 to the system resulted in a tight compaction of the dinucleosomal structure.

Animals↗

Longitudinal patterns similar to G-banding in untreated human chromosomes: evidence from atomic force microscopy.

The structure of human metaphase chromosomes, fixed according to standard procedures for optical microscopy but not treated for banding, was examined by atomic force microscopy (AFM). The images show that chromosomes display a banding pattern very similar to G-banding, detected by the AFM as a variation in the thickness of chromatin. This similarity allows the identification of individual chromosomes.

Chromatin↗

Atomic force microscopy studies of intercalation-induced changes in plasmid DNA tertiary structure.

Structural transitions in the tertiary structure of plasmid DNA have been investigated using atomic force microscopy. Changes in superhelical stress were induced by ethidium bromide intercalation, and conformational effects monitored by recording topographic images from DNA complexes of various ethidium bromide:base pair stoichiometry. Significant changes in the tertiary structure of individual DNA molecules were observed with increasing ethidium bromide concentration. The first distinct conformational transition was from a predominantly relaxed structure to one consisting solely of toroidal supercoils. A further increase in ethidium bromide concentration resulted in the formation of regions of plectonemic supercoiling. The ratio of plectonemic:toroidal supercoiling gradually increased until an extremely tightly interwound structure of solely plectonemic supercoiling was finally adopted. The toroidal form of supercoiling observed in this study is unusual as both atomic force microscopy and electron microscopy techniques have previously shown that plectonemic supercoiling is the predominant form adopted by plasmid DNA.

DNA, Superhelical↗

The effect of raft lipid depletion on microvilli formation in MDCK cells, visualized by atomic force microscopy.

We have investigated whether raft lipids of Madin-Darby canine kidney (MDCK) cells play any role in microvilli maintenance using a combination of atomic force microscopy (AFM) and laser scanning confocal microscopy. MDCK cells were treated to reduce the amount of sphingolipids, cholesterol, or both and subsequently imaged, in buffer solution, using AFM. It was observed that inhibition of either sphingolipid or cholesterol biosynthesis led to a reduction in the number of microvilli on the surface of MDCK cells. However, this effect was not uniform across the monolayer, with some cells resembling those in untreated controls. The subsequent extraction of cholesterol from cells grown in the presence of inhibitors led to a further reduction in microvilli on the surface of the cells and, in some cases, resulted in monolayers devoid of full length microvilli. Significantly, smaller spikes were observed on the surface of the smoother cells.

Actins↗