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At least 811 records · Page 45Linked to original sources
PubMed1998

Molecular structure of human topoisomerase II alpha revealed by atomic force microscopy.

The entire human topoisomerase II alpha (hTopoII alpha) dimer was expressed in the yeast Saccaromyces cerevisiae, purified to homogeneity, and subjected to atomic force microscopy (AFM) under a tapping mode. Molecular images obtained exhibited a 'heart or donut-like' structure with a large axial hole. The main benefit of the application of AFM to study the hTopoII alpha is that clear images of the internal 'pore' have been achieved without crystallization, staining, or fixation of the sample. These images are consistent with the model in which topoisomerase II has a large internal gate for DNA strand passage.

Antigens, Neoplasm↗
PubMedDate not supplied

Cantilevers and tips for atomic force microscopy.

The cantilever and the tip are at the centerpiece of the AFM. Properties such as the cantilever stiffness and resonant frequency, tip shape and sharpness, and material characteristics determine the mode of operation of the AFM and the type of experiments and measurements that can be performed. The possibility of batch fabricating cantilevers has permitted the fabrication and characterization of specialized tips for a variety of experiments. We believe that the use of new materials and tip shapes will allow new applications for the AFM in the future.

Aluminum↗
PubMed2005

The stepwise process of chromium-induced DNA breakage: characterization by electrochemistry, atomic force microscopy, and DNA electrophoresis.

DNA conformational change and breakage induced by Cr(VI)-GSH interaction were characterized by the integrated tools of electrochemistry, atomic force microscopy (AFM), and DNA electrophoresis. While electrochemistry confirmed the formation of the active species generated from Cr(VI)-GSH reduction, which causes the DNA conformational changes, AFM imaging vividly demonstrated the stepwise process of the DNA denaturation and breakage for the first time. Our DNA electrophoresis further validated that the DNA breakage occurs unevenly at both of the single strands of the molecule. A scheme was drawn based on the experimental observations to explain the phenomenon of the Cr-induced DNA cleavage.

Animals↗
PubMedDate not supplied

[Use of atomic force microscopy to study the morphology and structure of viruses].

The paper describes AFM methodic approaches in the investigation of viruses (TMV, phage T4). A possibility is shown to use this method to obtain high-quality three-dimensional image of objects without additional treatment with salts of heavy metals. The phage T4 was also a model to study the object density, using the phase shift of the AFM. Such researches can determine the internal structure of a virion and folding of nucleic acid inside a virion.

Bacteriophage T4↗
PubMed2001

The assembly of amyloidogenic yeast sup35 as assessed by scanning (atomic) force microscopy: an analogy to linear colloidal aggregation?

Amyloidosis is a class of diseases caused by protein aggregation and deposition in various tissues and organs. In this paper, a yeast amyloid-forming protein Sup35 was used as a model for understanding amyloid fiber formation. The dynamics of amyloid formation by Sup35 were studied with scanning force microscopy. We found that: 1) the assembly of Sup35 fibers begins with individual NM peptides that aggregate to form large beads or nucleation units which, in turn, form dimers, trimers, tetramers and longer linear assemblies appearing as a string of beads; 2) the morphology of the linear assemblies differ; and 3) fiber assembly suggests an analogy to the aggregation of colloidal particles. A dipole assembly model is proposed based on this analogy that will allow further experimental testing.

Amyloid↗
PubMed2002

Supported lipid bilayers as effective substrates for atomic force microscopy.

Supported lipid bilayers offer a diverse set of substrates for AFM investigations of both water-soluble samples and integral membrane proteins. Although their amphipathic nature is necessary for the latter specimens, it is their robustness and easily changed surface characteristics that make these surfaces particularly attractive for the former ones. When starting any AFM investigation of water-soluble biological complexes, it is probably best to try mica first, owing to its remarkable effectiveness with a wide range of samples. Yet, if mica should prove inadequate, supported lipid bilayers are, as demonstrated here, a sensible second choice.

Aluminum Silicates↗
PubMed2002

Ripples and the formation of anisotropic lipid domains: imaging two-component supported double bilayers by atomic force microscopy.

Direct visualization of the fluid-phase/ordered-phase domain structure in mica-supported bilayers composed of 1,2-dimyristoyl-sn-glycero-3-phosphocholine/1,2-distearoyl-sn-glycero-3-phosphocholine mixtures is performed with atomic force microscopy. The system studied is a double bilayer supported on a mica surface in which the top bilayer (which is not in direct contact with the mica) is visualized as a function of temperature. Because the top bilayer is not as restricted by the interactions with the surface as single supported bilayers, its behavior is more similar to a free-standing bilayer. Intriguing straight-edged anisotropic fluid-phase domains were observed in the fluid-phase/ordered-phase coexistence temperature range, which resemble the fluid-phase/ordered-phase domain patterns observed in giant unilamellar vesicles composed of such phospholipid mixtures. With the high resolution provided by atomic force microscopy, we investigated the origin of these anisotropic lipid domain patterns, and found that ripple phase formation is directly responsible for the anisotropic nature of these domains. The nucleation and growth of fluid-phase domains are found to be directed by the presence of ripples. In particular, the fluid-phase domains elongate parallel to the ripples. The results show that ripple phase formation may have implications for domain formation in biological systems.

Anisotropy↗
PubMed1999

The structure of the nucleosome core particle of chromatin in chicken erythrocytes visualized by using atomic force microscopy.

The structure of the nucleosome core particle of chromatin in chicken erythrocytes has been examined by using AFM. The 146 bp of DNA wrapped twice around the core histone octamer are clearly visualized. Both the ends of entry/exit of linker DNA are also demonstrated. The dimension of the nucleosome core particles is approximately 1-4 nm in height and approximately 13-22 nm in width. In addition, superbeads (width of approximately 48-57 nm, height of approximately 2-3 nm) are occasionally revealed, two turns of DNA around the core particles are also detected.

Animals↗
PubMed2006

Amyloid fibrils of glucagon characterized by high-resolution atomic force microscopy.

Glucagon solutions at pH 2.0 were subjected to mechanical agitation at 37 degrees C in the presence of a hydrophobic surface to explore the details of aggregation and fiber formation. High-resolution intermittent-contact atomic force microscopy performed in solution revealed the presence of aggregates after 0.5 h; however, longer agitation times resulted in the formation of fibrillated structures with varying levels of higher-order assembly. Height, periodicity, and amplitude measurements of these structures allowed the identification of four distinct fiber types. The most elementary fiber form, designated a filament, self-associates in a specific wound fashion to produce protofibrils composed of two filaments. Subsequent self-assembly of these filaments and protofibrils leads to two well-defined fibrillar motifs, termed Type I and Type II. Atomic force microscopy imaging of pH 2.8 glucagon solutions not agitated or exposed to elevated temperature revealed the presence of amorphous aggregates before the formation of fibrillar structures similar to those seen at pH 2.0. Time-course solution Fourier transform infrared spectroscopy and thioflavin T binding studies suggested that glucagon aggregation and fibril formation were associated with the development of beta-sheet structure. The results of these studies are used to describe a possible mechanism for glucagon aggregation and fibrillation that is consistent with a hierarchical assembly model proposed for amyloid fibril formation.

Amyloid↗
PubMed2005

Effects of 2% chlorhexidine and 5.25% sodium hypochlorite on gutta-percha cones studied by atomic force microscopy.

AIM: To compare the effects of 2% chlorhexidine (CHX) and 5.25% sodium hypochlorite (NaOCl) on gutta-percha (GP) cone structure using atomic force microscopy (AFM). METHODOLOGY: Two standardized GP cones were sectioned 3 mm from the tip, attached to a glass base and immersed in 2% CHX or 5.25% NaOCl for 1, 5, 10, 20 and 30 min. Untreated GP cones were used as control. Topography and elasticity analyses were performed on 12 different regions located between 1 and 2 mm from the tip. Root mean square (RMS) parameters for contact mode imaging and force modulation microscopy variations were measured. The differences between RMS values were tested by anova with Fisher's protected LSD test for multiple comparisons. RESULTS: There was no deterioration in the topography and physical properties studied when 2% CHX was used in comparison with the control (P < 0.05). The RMS parameter for topography increased after 10 min of 5.25% NaOCl exposure in comparison with the control (P < 0.05). In addition, 5.25% NaOCl increased the elasticity of the GP cone after an immersion time of 1 min in comparison with the control (P < 0.05). CONCLUSIONS: Two per cent CHX did not change GP cone structure following up to 30 min exposure. Conversely, 5.25% NaOCl caused elastic changes after 1 min exposure.

Chlorhexidine↗
PubMed2002

Daptomycin morphostructural damage in Bacillus cereus visualized by atomic force microscopy.

Daptomycin is a novel, rapidly bactericidal in vitro antibiotic that is under investigation for the treatment of serious Gram-positive infections. Although daptomycin appears to disrupt membrane function, the precise mechanism of action has not been fully elucidated. Atomic force microscopy (AFM) is an innovative technique that allows high-resolution visualization and digital image manipulation of cell surface structures in 3 dimensions without the use of photons and electrons. The aim of this study was to use AFM to investigate the morphostructural changes in Bacillus cereus that occur upon daptomycin administration. The effects of daptomycin at 4x and 8x the minimal inhibitory concentration were visualized during an 8-hour incubation period. Atomic force microscopy images showed aberrant bacterial surface formations, including flattening and shrinking of cells and leakage of cytoplasm through the membrane. In addition to structural changes, the destabilization of flagella was also observed. These results support previous data suggesting that daptomycin disrupts membrane function.

Anti-Bacterial Agents↗
PubMed2004

Atomic force microscopy characterization of an electrochemical DNA-biosensor.

Electrode surface characteristics represent an important aspect on the construction of sensitive DNA electrochemical biosensors for rapid detection of DNA interaction and damage. Two different immobilization procedures of double-stranded DNA (dsDNA) at the surface of a HOPG electrode were evaluated by MAC mode AFM performed in air. A thin dsDNA adsorbed film forming a network structure with holes exposing the electrode surface and a thick dsDNA film completely covering the electrode surface, presenting a much rougher structure, were investigated. The DNA surface characteristics and structure are discussed with respect to the degree of surface coverage.

Adsorption↗
PubMed1995

Immunolocalization of lamins and nuclear pore complex proteins by atomic force microscopy.

The nuclear envelope functions as a selective barrier separating the nuclear from the cytosolic compartment. Nuclear pore complexes (NPCs) mediate nuclear import and export of macromolecules and, therefore, are potential regulators of gene expression. In this study we applied atomic force microscopy (AFM) to visualize the three dimensional (3D) structure of individual NPCs in the absence and presence of two different antibodies, one directed against a pore protein (gp62) and another directed against Xenopus lamin LIII, a component of the nuclear lamina, a filament meshwork localized on the nucleoplasmic side of the nuclear envelope (NE) adjacent to and interacting with NPCs. Using 12-nm gold-labelled secondary antibodies and transmission electron microscopy we could clearly localize the primary single anti-gp62 antibody on NPCs and the primary single anti-LIII antibody between NPCs. Using AFM, the secondary antibodies against anti-gp62 could be detected as particles 7 nm in height on the nucleoplasmic face of NPCs. The secondary antibodies against anti-LIII could be clearly identified between NPCs. The secondary antibodies, attached to a 12-nm colloidal gold particle and visualized on glass, revealed similar shapes and heights as found on NEs. According to the 3D images, the volume of a single gold particle conjugated with secondary antibodies was 10203 nm3. This volume is equivalent to the volume of 38 IgG molecules associated with one individual gold particle. A similar volume of 11987 nm3 was calculated from a model assuming that the 150-kDa IgG molecules perfectly cover the spherical gold particle. We conclude that AFM can be used for identifying antibodies or other macromolecules associated with biomembranes.

Animals↗
PubMed2005

Predation, death, and survival in a biofilm: Bdellovibrio investigated by atomic force microscopy.

Biofilms are complex microbial communities that are resistant to attack by bacteriophages and to removal by drugs and chemicals. Here we use atomic force microscopy (AFM) to image the attack on Escherichia coli biofilms by Bdellovibrio bacteriovorus 109J. Bdellovibrio is a small, predatory bacterium that invades and devours other Gram-negative bacteria. We demonstrate that under dilute nutrient conditions, bdellovibrios can prevent the formation of simple bacterial biofilms and destroy established biofilms; under richer conditions the prey bacteria persist and are not eradicated, but may be shifted toward solution populations. Using AFM we explore these bacterial interactions with more detail and accuracy than available by more traditional staining assays or optical microscopy. AFM also allows us to investigate the nanoscale morphological changes of the predator, especially those related to motility. This demonstration of Bdellovibrio's successful predation in a biofilm inspires us to consider ways that it might be used productively for industrial, medical, agricultural, and biodefensive purposes.

Adhesiveness↗
PubMed2004

Investigation of the molecular interactions in a pMDI formulation by atomic force microscopy.

The forces of interaction between inhalable formoterol fumarate dihydrate particles, and the various components of a pressurised Metered Dose Inhaler (pMDI) (e.g. aluminium container, other drug particles and stabilising excipients) were investigated in isolation and combination in the model propellant 2H, 3H perfluoropentane (HPFP). The results obtained offer conclusive proof that the model systems tested have a degree of instability both in the presence and absence of polymer(s). The attractive forces measured following the addition of a mixed homopolymer solution may well be weak enough to be overcome by shaking. The use of homopolymers has inherent difficulties associated with it in the form of bridging interaction. Block or comb copolymers may well be a better option therefore, as they will reduce the possibility of bridging interactions. AFM offers great insight into the behaviour of HFA drug suspensions, and could play a vital role in the future development of suspension formulations.

Bronchodilator Agents↗
PubMed2006

Relationship between apical membrane elasticity and stress fiber organization in fibroblasts analyzed by fluorescence and atomic force microscopy.

To investigate the relationship between cellular microelasticity and the structural features of cytoskeletons (CSKs), a microindentation test for apical cell membranes and observation of the spatio-distribution of actin CSKs of fibroblasts were performed by fluorescence and atomic force microscopy (FM/AFM). The indentation depths of apical cell membranes were measured from AFM force-indentation (f-i) curves under equal final loads and mapped two-dimensionally to show the relative distribution of local microelasticity on cell membranes. Intracellular spatial distribution of actin CSKs was visualized fluorescently by high Z-resolution cross-sectional observation of a cell on which indentation mapping analysis had been performed in advance. Structural features of stress fibers (SFs) were observed as three typical patterns of dense SF, sparse SF and sparser SF cell groups, which were quantitated using the degree of orientation in apical SFs (ASFs) that had been defined using two-dimensional Fourier analysis. In indentation depth maps, the upper nuclear region was markedly softer than the pseudopodium region. The mean indentation depth of the upper nuclear region decreased with increased SF density in whole cells and the degree of orientation of ASF, although the pseudopodium region did not exhibit such a trend. The apical membrane of adhered cells was found to tend to stiffen with the increase in both density and degree of orientation of SFs.

Actins↗
PubMed2002

Absolute measurement of phosphorylation levels in a biological membrane using atomic force microscopy: the creation of phosphorylation maps.

We show that it is possible to produce phosphorylation difference maps of biological membranes under conditions which reflect those in vivo and in which proteins remain functional. We also demonstrate that absolute levels of phosphorylation are retrieved through the application of an appropriate calibration method. Finally we show that the kinetics of phosphorylation/dephosphorylation can also be monitored. These methods are demonstrated on photosynthetic membranes from higher plants, for which protein phosphorylation is the dominant regulatory mechanism. We show directly that the most recent estimates of the phosphorylation levels in this system are reasonably accurate. Phosphorylation difference maps show that the distribution of phosphates is not even, with significantly higher levels at the membrane margins and patches of high phosphate density next to patches of low charge density.

Membranes↗
PubMed2004

Molecular weight and polydispersity estimation of adsorbing polymer brushes by atomic force microscopy.

We have estimated the molecular weight, Mn, and polydispersity, PDI, of densely grafted poly(N-isopropylacrylamide) (PNIPAM) brushes using a novel atomic force microscopy (AFM) approach. When compression of a polymer brush induced adsorption of multiple chains to an AFM tip, the resulting decompression force profile exhibited a maximum attractive force at a separation, Lm, that decayed to zero with increasing tip-sample separation. We have found that the separation Lm approximates the average contour length, Lc, determined by gel permeation chromatography (GPC). The detection of a decaying attractive force at separations larger than Lc suggests that chains of above average length sequentially break free from the tip as they are stretched away from the grafting surface. The shape of the decompression profile in this region approximately paralleled the cumulative weight fraction of the grafted chains determined by GPC. The fraction of chains of a given molecular weight determined from a single force curve fit a log-normal distribution, having a standard deviation that provided an estimate of the PDI. We have characterized two PNIPAM brushes by this AFM technique as well as by GPC coupled to a multiangle laser light-scattering detector (MALLS). The values obtained by AFM-(1) Mn,AFM = (3.8+/-0.5) x 10(4), PDI,(AFM) = 1.3+/-0.1 and (2) Mn,AFM = (9.4+/-1.4) x 10(4), PDI,(AFM) = 1.3+/-0.1-agreed quite well with the corresponding GPC/MALLS values of (1) Mn,GPC = 4.77 x 10(4), PDI,GPC = 1.33 and (2) Mn,GPC = 9.49 x 10(4), PDI = 1.35. This technique requires only a single force curve to obtain a statistical distribution of contour lengths and provides a novel method for estimating the Mn and PDI of appropriate uniformly grafted dense polymer layers.

Acrylic Resins↗