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At least 235 records · Page 13Linked to original sources
PubMed2000

MutS-mediated detection of DNA mismatches using atomic force microscopy.

We have developed an atomic force microscopy-based method for detecting DNA base-pair mismatches using MutS protein isolated from E. coli. MutS is a biological sensor and a locator of DNA base-pair mismatches. It binds specifically to a mismatched DNA base pair and initiates a process of DNA repair. To test the possibility of visually detecting mismatched base pairs by atomic force microscopy, we prepared DNA templates approximately 500 bp in length consisting of a single or multiple base-pair mismatches. We demonstrate that MutS binding sites on individual DNA molecules were readily detectable by atomic force microscopy and that the observed positions were in good agreement with the predicted sites of base-pair mismatches at a few-nanometer resolution. The technique described here is rapid and sensitive and is expected to be useful in screening mutations and DNA polymorphisms.

Adenosine Triphosphatases↗
PubMed2004

Effects of acetic acid treatment on plant chromosome structures analyzed by atomic force microscopy.

Acetic acid treatment has been frequently used to remove cellular contaminants from plant chromosome samples for structural analyses by scanning electron microscopy and atomic force microscopy (AFM). We evaluated the effects of various concentrations of acetic acid treatments on barley chromosome structures by using AFM. The long-term 45% acetic acid treatment significantly damaged the chromosome structures, although the treatment effectively removed the cellular contaminants. On the other hand, the treatment with 15% acetic acid could not obtain sufficiently clean chromosome samples and the chromosome surface structures could not be observed. In contrast, we obtained clean chromosome preparation without severe damage by using an intermediate concentration (30%) of acetic acid treatment. In the centromeric region, we could observe fiber structures with a width of 100 nm, which were composed of ca. 50-nm granules and aligned to the axes of chromosomes. Thus, AFM analysis of chromosomes appropriately treated with acetic acid will provide important insights into the organization of higher-order structures of plant chromosomes.

Acetic Acid↗
PubMed2007

Synthesis and in situ atomic force microscopy characterization of temperature-responsive hydrogels based on poly(2-(dimethylamino)ethyl methacrylate) prepared by atom transfer radical polymerization.

Well-defined copolymers of 2-(dimethylamino)ethyl methacrylate (DMAEMA) and benzophenone methacrylate (BPMA) with different compositions were synthesized via atom transfer radical polymerization. The molecular weights of these copolymers were Mn approximately 30 000 g/mol, while the BPMA content varied from 2.5 to 10 mol %. The copolymers with a low content of BPMA (2.5 and 5 mol %) exhibited a sharp thermal transition at 33-36 degrees C in aqueous solution. A hydrogel was immobilized and patterned on a silicon wafer via UV treatment of the spin-coated polymer layer using a photomask technique. The thermoresponsive behavior of the patterned polymer gel was quantitatively investigated by variable temperature in situ contact mode atomic force microscopy, which revealed the presence of two lower critical solution temperature regions. One region was between 25 and 30 degrees C, corresponding to the topmost layer of the hydrogel film, and the other region, around 40 degrees C, corresponded to the bulk of the hydrogel. Concurrent lateral force microscopy measurements revealed that, just above the transition temperature, the bulk region exhibited enhanced friction.

Free Radicals↗
PubMed2000

The application of atomic force microscopy to topographical studies and force measurements on the secreted adhesive of the green alga Enteromorpha.

Atomic force microscopy (AFM) enables the topographical structure of cells and biological materials to be resolved under natural (physiological) conditions, without fixation and dehydration artefacts associated with imaging methods in vacuo. It also provides a means of measuring interaction forces and the mechanical properties of biomaterials. In the present study, AFM has been applied for the first time to the study of the mechanical properties of a natural adhesive produced by a green plant cell. Swimming spores of the green alga Enteromorpha linza (L.) J. Ag. (7-10 microm) secrete an adhesive glycoprotein which provides firm anchorage to the substratum. Imaging of the adhesive in its hydrated state revealed a swollen gel-like pad, approximately 1 microm thick, surrounding the spore body. Force measurements revealed that freshly released adhesive has an adhesion strength of 173 +/- 1.7 mN m(-1) (mean +/- SE; n=90) with a maximum value for a single adhesion force curve of 458 mN m(-1). The adhesive had a compressibility (equivalent to Young's modulus) of 0.54 x 10(6) +/- 0.05 x 10(6) N m-2 (mean +/- SE; n=30). Within minutes of release the adhesive underwent a progressive 'curing' process with a 65% reduction in mean adhesive strength within an hour of settlement, which was also reflected in a reduction in the average length of the adhesive polymer strands (polymer extension) and a 10-fold increase in Young's modulus. Measurements on the spore surface itself revealed considerably lower adhesion-strength values but higher polymer-extension values than the adhesive pad, which may reflect the deposition of different polymers on this surface as a new cell wall is formed. The study demonstrates the value of AFM to the imaging of plant cells in the absence of fixation and dehydration artefacts and to the characterisation of the mechanical properties of plant glycoproteins that have potential utility as adhesives.

Cell Adhesion↗
PubMed2001

Imaging erythrocytes under physiological conditions by atomic force microscopy.

Since its invention in the mid 1980s atomic force microscopy has revolutionised the way in which surfaces can be imaged. Close to atomic resolution has been achieved for some materials and numerous images of molecules on surfaces have been recorded. Atomic force microscopy has also been of benefit to biology where protein molecules on surfaces have been studied and even whole cells have been investigated. Here we report a study of red blood cells which have been imaged in a physiological medium. At high resolution, the underlying cytoskeleton of the blood cell has been resolved and flaws in the cytoskeleton structure may be observed. Comparison of the normal 'doughnut' shaped cells with swollen cells has been undertaken. Differences in both the global properties of the cells and in the local features in cytoskeleton structure have been observed.

Animals↗
PubMed2006

Controlled loading of oligodeoxyribonucleotide monolayers onto unoxidized crystalline silicon; fluorescence-based determination of the surface coverage and of the hybridization efficiency; parallel imaging of the process by Atomic Force Microscopy.

Unoxidized crystalline silicon, characterized by high purity, high homogeneity, sturdiness and an atomically flat surface, offers many advantages for the construction of electronic miniaturized biosensor arrays upon attachment of biomolecules (DNA, proteins or small organic compounds). This allows to study the incidence of molecular interactions through the simultaneous analysis, within a single experiment, of a number of samples containing small quantities of potential targets, in the presence of thousands of variables. A simple, accurate and robust methodology was established and is here presented, for the assembling of DNA sensors on the unoxidized, crystalline Si(100) surface, by loading controlled amounts of a monolayer DNA-probe through a two-step procedure. At first a monolayer of a spacer molecule, such as 10-undecynoic acid, was deposited, under optimized conditions, via controlled cathodic electrografting, then a synthetic DNA-probe was anchored to it, through amidation in aqueous solution. The surface coverage of several DNA-probes and the control of their efficiency in recognizing a complementary target-DNA upon hybridization were evaluated by fluorescence measurements. The whole process was also monitored in parallel by Atomic Force Microscopy (AFM).

Biosensing Techniques↗
PubMed2006

Atomic-scale sharpening of silicon tips in noncontact atomic force microscopy.

The atomic-scale stability of clean silicon tips used in noncontact atomic force microscopy (NC-AFM) is simulated by ab initio calculations based on density functional theory. The tip structures are modeled by silicon clusters with and termination. For the often assumed Si(111)-type tip we observe the sharpening of the initially blunt tip via short-range chemical forces during the first approach and retraction cycle. The structural changes corresponding to this intrinsic process are irreversible and lead to stable NC-AFM imaging conditions. In opposition to the picture used in literature, the Si(001)-type tip does not exhibit the so-called "two-dangling bond" feature as a bulklike termination suggests.

Journal Article↗
PubMed1999

[Study of erythrocytes by atomic force microscopy].

Working principle of Nanotop-202 atomic force microscope is described. The morphology of peripheral blood erythrocytes is studied under this microscope. A method for preparing the specimens is developed and the resultant cell images are described. The results demonstrate the potentialities of the method for morphometry, studies of fine structure of membrane surface, and manipulations with erythrocytes. Prospects of atomic force microscopy in hematological studies are discussed.

Animals↗
PubMed2004

On atomic force microscopy and the constitutive behavior of living cells.

Atomic force microscopy (AFM) is one of many new technologies available to study the mechanical properties and mechanobiological responses of living cells. Despite the widespread usage of this technology, there has been little attempt to develop new theoretical frameworks to interpret the associated data. Rather, most analyses rely on the classical Hertz solution for the indentation of an elastic half-space within the context of linearized elasticity. In contrast, we propose a fully nonlinear, constrained mixture model for adherent cells that allows one to account separately for the contributions of the three primary structural constituents of the cytoskeleton. Moreover, we extend a prior solution for a small indentation superimposed on a finite equibiaxial extension by incorporating in this mixture model for the special case of an initially random distribution of constituents (actin, intermediate filaments, and microtubules). We submit that this theoretical framework will allow an improved interpretation of indentation force-depth data from a sub-class of atomic force microscopy tests and will serve as an important analytical check for future finite element models. The latter will be necessary to exploit further the capabilities of both atomic force microscopy and nonlinear mixture theories for cell behavior.

Biomechanical Phenomena↗
PubMed2003

Single complexation force of 18-crown-6 with ammonium ion evaluated by atomic force microscopy.

The cation complexation force of 18-crown-6 has been measured in ethanol by means of atomic force microscopy using probe tips and mica substrates modified chemically with 18-crown-6 and ammonium ion, respectively. The specific complexation force was suppressed by free potassium ion in the measurement solution, indicating a blocking effect based on the competitive complexation of the 18-crown-6 moiety between the free ion and the ammonium ion bound to the substrate. The single complexation force of 18-crown-6 with ammonium ion was evaluated to be about 60 pN in ethanol by autocorrelation analyses of the histograms for observed forces.

Crown Ethers↗
PubMed2000

Optical and atomic force microscopy of an explanted AcrySof intraocular lens with glistenings.

PURPOSE: To assess the surface morphology and cause of glistenings in an explanted AcrySof intraocular lens (IOL). SETTING: Shakai Hoken Kobe Central Hospital, Kobe, Japan. METHODS: A 63-year-old Japanese man had implantation of an AcrySof IOL in the capsular bag. One month postoperatively, he had a neodymium:YAG laser capsulotomy for posterior capsule opacification, which changed the IOL's position in the capsular bag. A few months later, the patient developed disabling night glare from intralenticular glistenings and progressive hyperopic refractive error. The IOL was explanted and then analyzed by optical microscopy and atomic force microscopy (AFM). Laboratory analysis of control AcrySof IOLs kept in a balanced salt solution at steady room and body temperature for 2 months was also performed to evaluate the cause of the glistenings observed clinically. RESULTS: Optical microscopy showed that the explanted AcrySof IOL had several microvacuoles; no abnormalities were observed in the control AcrySof IOLs before or after folding at the room and body temperatures. The AFM analysis showed a significant change in the surface morphology of the explanted IOL, including vacuolar formations in the posterior surface as well as numerous anterior surface irregularities. No microvacuoles or surface morphology alterations were observed in the control AcrySof IOLs by AFM analysis. CONCLUSIONS: The glistenings in the explanted AcrySof IOL were likely caused by temperature changes and not mechanical stress from folding.

Acrylic Resins↗
PubMed2001

Detection of abasic sites on individual DNA molecules using atomic force microscopy.

We have developed an atomic force microscopy-based method for detecting abasic sites (AP sites) on individual DNA molecules. By using uracil and uracil DNA glycosylase, we first prepared a 250-bp DNA template consisting of two AP sites at specific locations. We then detected the AP sites by marking them with biotinylated aldehyde-reactive probes and monomeric avidin. We demonstrate here that (i) the location of monomeric avidin bound on a single DNA molecule was detectable by atomic force microscopy; (ii) the observed location of avidin was in good agreement to the predicted AP sites at a few nanometer resolution; and (iii) by end-labeling the 5'-terminus of one DNA strand, the AP sites were determined without directional ambiguity. The technique described here will provide a sensitive way of locating AP sites and contribute to screen DNA damages from individual molecules.

DNA↗
PubMed1999

Collagen II containing a Cys substitution for Arg-alpha1-519. Analysis by atomic force microscopy demonstrates that mutated monomers alter the topography of the surface of collagen II fibrils.

A recombinant human procollagen II was prepared that contained a substitution of Cys for Arg at alpha1-519 and that was found in five families with early onset generalized osteoarthritis with or without features of a mild chondrodysplasia. Previously, the presence of mutated monomers in mixtures with wildtype collagen II was shown to increase the lag period for fibril assembly. Also, the fibrils were more loosely packed and some thick fibrils lacked a D-periodic banding pattern. Here we re-examined the fibrils using a combination of transmission electron microscopy and atomic force microscopy. The presence of the mutated monomers increased the diameter of the thin filaments that were consistently formed in association with the thick fibrils of collagen II. In addition, the presence of the mutated monomers increased the depth of the gap regions in all fibrils with a distinct D-periodic banding pattern. The results, therefore, may indicate that the mutated monomers formed two or three additional outer layers of monomers in 0D-period staggers on the surface of the fibrils. Apparently, the mutated monomers were bound on the surface through intermolecular disulfide bonds.

Amino Acid Substitution↗
PubMed2006

Imaging by atomic force microscopy of the plasma membrane of prestin-transfected Chinese hamster ovary cells.

The high sensitivity of mammalian hearing is achieved by amplification of the motion of the cochlear partition. This cochlear amplification is thought to be generated by the elongation and contraction of outer hair cells (OHCs) in response to acoustical stimulation. This motility is made possible by a membrane protein embedded in the lateral membrane of OHCs. Although a fructose transporter, GLUT-5, was initially proposed to be this protein, a later study identified the gene of the motor protein distributed throughout the OHC plasma membrane. This protein has been named "prestin." However, although previous morphological studies by electron microscopy and atomic force microscopy (AFM) found the lateral wall of OHCs to be covered with 10-nm particles, believed to be motor proteins, it is unknown whether such particles consist only of prestin or are a complex of GLUT-5 and prestin molecules. To determine if the 10-nm particles are indeed constituted only of prestin, plasma membranes of prestin-transfected and untransfected Chinese hamster ovary (CHO) cells, which do not express GLUT-5, were observed by AFM. First, the cells attached to a substrate were sonicated so that only the plasma membrane remained on the substrate. The cytoplasmic face of the cell was observed by the tapping mode of the AFM in liquid. As a result, particle-like structures were recognized on the plasma membranes of both the prestin-transfected and untransfected CHO cells. Comparison of the difference in the frequency distribution of these structures between those two cells showed approximately 75% of the particle-like structures with a diameter of 8-12 nm in the prestin-transfected CHO cells to be possibly constituted only by prestin molecules. Our data suggest that the densely packed 10-nm particles observed on the OHC lateral wall are likely to be constituted only of prestin molecules.

Animals↗
PubMed2004

3-D morphological characterization of the liver parenchyma by atomic force microscopy and by scanning electron microscopy.

A comparative study of atomic force microscopy (AFM) and scanning electron microscopy (SEM) imaging of the healthy human liver parenchyma was carried out to determine the similarities and the differences. In this study, we compared the fine hepatic structures as observed by SEM and AFM. Although AFM revealed such typical hepatic structures as bile canaliculi and hepatocytes, it also showed the location of the nucleus and chromatin granules in rough relief structure, which was not visible by SEM. By contrast, SEM visualized other structures, such as microvilli, the central vein, and collagenous fibers, none of which was visualized by AFM. For better orientation and confirmation of most of the structures imaged by SEM and AFM, Congo Red-stained specimens were also examined. Amyloid deposits in the Disse's spaces were shown especially clearly in these images. The differences between the SEM and AFM images reflected the characteristics of the detection systems and methods used for sample preparation. Our results reveal that more detailed information on hepatic morphology is obtained by exploiting the advantages of both SEM and AFM.

Adult↗
PubMed2005

Three-dimensional morphological characterization of optic nerve fibers by atomic force microscopy and by scanning electron microscopy.

A comparative study of scanning electron microscopy (SEM) and atomic force microscopy (AFM) imaging of the healthy human optic nerve was carried out to determine the similarities and the differences. In this study we compared the fine optic nerve structures as observed by SEM and AFM. The fibers of the right optic nerve of a 61-year-old man show different arrangements in transverse sections taken from the same individual 5 mm central to the optic canal and 5 mm peripheral to the optic chiasma; this difference can be recognized by light microscopy (LM), SEM, and AFM. AFM revealed such typical optic nerve fibers (taken from a point 5 mm central to the optic canal) with annular and longitudinal orientations, which were not visible by SEM in this form. By contrast, LM and SEM visualized other structures, such as pia mater and optic nerve fibers loosely arranged in bundles, none of which was visualized by AFM. The images, however, taken 5 mm peripheral from the optic chiasma show shapeless nerve fibers having a wavy course. Our results reveal that more detailed information on optic nerve morphology is obtained by exploiting the advantages of both SEM and AFM. These are the first SEM and AFM images of healthy human optic nerve fibers, containing clear representations of the three dimensions of the optic nerve.

Aged↗
PubMed2006

Energy dissipation and dynamic response of an amplitude-modulation atomic-force microscopy subjected to a tip-sample viscous force.

In a common environment of atomic force microscopy (AFM), a damping force occurs between a tip and a sample. The influence of damping on the dynamic response of a cantilever must be significant. Moreover, accurate theory is very helpful for the interpretation of a sample's topography and properties. In this study, the effects of damping and nonlinear interatomic tip-sample forces on the dynamic response of an amplitude-formulation AFM are investigated. The damping force is simulated by using the conventional Kelvin-Voigt damping model. The interatomic tip-sample force is the attractive van der Waals force. For consistance with real measurement of a cantilever, the mathematical equations of the beam theory of an AM-AFM are built and its analytical solution is derived. Moreover, an AFM system is also simplified into a mass-spring-damper model. Its exact solution is simple and intuitive. Several relations among the damping ratio, the response ratio, the frequency shift, the energy dissipation and the Q-factor are revealed. It is found that the resonant frequencies and the phase angles determined by the two models are almost same. Significant differences in the resonant quality factors and the response ratios determined by using the two models are also found. Finally, the influences of the variations of several parameters on the error of measuring a sample's topography are investigated.

Microscopy, Atomic Force↗