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PubMed2002

Binding of dentin noncollagenous matrix proteins to biological mineral crystals: an atomic force microscopy study.

Noncollagenous matrix proteins (NCPs) of dental hard tissues (dentin, cementum) are involved, both temporally and spatially, in the mineralization of their collagen matrices. Two of the NCPs thought to initiate mineral nucleation and control crystal growth in dentin, are dentin phosphoproteins (DPP) and dentin sialoprotein (DSP). Control of crystal growth would depend on the binding capacity of these two molecules, which may be related to the charge domains on the crystals and/or the phosphorylation of the protein. Phosphophoryn (a highly phosphorylated DPP) and DSP were isolated, purified, and characterized from the immature root apicies of human teeth. Dephosphorylation of phosphophoryn was carried out using bovine intestinal alkaline phosphatase. Enamel crystals were prepared from the maturation stage of developing rat incisor enamel. Protein-coated crystals were prepared for viewing in an atomic force microscope fluid cell using tapping mode. Desorption of the proteins was achieved using a phosphate buffer and surface roughness measurements were obtained from all specimens. Time-lapsed images of the crystals showed "nanospheres" of protein distributed along the crystals but only the phosphophoryn-coated crystals showed a distinctive banding pattern, which was still visible after the phosphate desorption experiments. The surface roughness measurements were statistically greater (P <0.01) when compared to the control for only the phosphophoryn-coated specimens. It is hypothesized that the phosphophoryn binding may be associated with charge arrays on the crystal surface and its phosphorylation. Also, based on its affinity for the crystalsurfaces, phosphophoryn seems the most likely candidate for controlling dentin crystal growth and morphology.

Crystallization↗
PubMed2006

Interfacial membrane effects of fluoroquinolones as revealed by a combination of fluorescence binding experiments and atomic force microscopy observations.

6-Fluoroquinolones are useful antimicrobial agents against gram-positive and gram-negative bacteria and some mycobacterial species as well. Although the diffusion through porins in gram-negative bacteria is well established, other mechanisms such as the hydrophobic pathway through the apolar regions of the bilayer and the self-promoted pathway appear to be relevant or concomitant with the hydrophilic pathway in many cases. This article discusses the interaction of ciprofloxacin (CPX) and two new synthesized compounds (M3CPX and M4CPX)-with a methyl group attached at the N3 and N4 positions of the piperazynil ring of the CPX-with liposomes and supported planar bilayers (SPBs) of Escherichia coli. Binding experiments using ANS revealed that the three compounds interact electrostatically with the bilayer. The variations in the electrostatic surface potential, which is always positive, were higher for M3CPX than for CPX or M4CPX. Related to that, the SPBs of E. coli were more affected by M3CPX than by the other two compounds, as judged by the analysis of the atomic force microcopy (AFM) images. The in situ injection of the three 6-fluoroquinolones (6-FQs) induced different changes in height, roughness (Ra), and area covered by the SPBs.

Escherichia coli↗
PubMed1999

Self-aggregation of DNA oligomers with XGG trinucleotide repeats: kinetic and atomic force microscopy measurements.

Turbidity measurements via absorbance monitoring at 320 nm were employed to obtain autocatalytic-like kinetic profiles of K+-induced aggregate formation of d(XGG)4 and some related oligomers, where X = A, C, G, and T. At least 1 M KCl is needed to observe the turbidity-measurable aggregation at pH 8, and the relative propensity for aggregate formation is shown to follow the order d(GGG)4 > d(AGG)4 approximately d(TGG)4 >> d(CGG)4. The presence of Mg2+ greatly facilitates and dramatically reduces the amount of K+ required to initiate aggregation and significantly enhances the thermal stabilities of the aggregates. Replacement of K+ by Na+ fails to induce a similar phenomenon. The Psi-type CD characteristics of aggregates are strongly dependent on the sequence and ionic conditions. Despite their ease of aggregate formation, oligomers with AGG trinucleotide repeats fail to exhibit Psi-CD formation. The propensity for aggregation is greatly affected by the chain length, with oligomers of four repeats being most facile. Appending X base at the 3' end of d(GGXGGXGGXGG) appears to provide a greater hindrance to aggregation than at the 5' end. Atomic force microscopic images support some of these findings and reveal the morphologies of these aggregates. The presence of MgCl2 in solutions appears to considerably elongate the K+-induced aggregates.

Circular Dichroism↗
PubMed2000

Observation of human corneal and scleral collagen fibrils by atomic force microscopy

Purpose: We attempted to analyze the three-dimensional ultrastructure of human corneal and scleral collagen fibrils with an atomic force microscope (AFM).Methods: A normal eye removed from a 66-year-old male was used in the study. Suspended corneal and scleral collagen fibrils were individually attached to glass slides by centrifugation. These collagen fibrils were air-dried and observed with a noncontact mode AFM in air.Results: AFM imaging provided information on the surface topography of both corneal and scleral collagen fibrils. The corneal collagen fibrils had a height of 11.9 +/- 1.0 (mean +/- standard deviation) nm and the scleral fibrils of 82.5 +/- 35.6 nm. A periodic banding pattern of grooves and ridges was clearly found in both types of fibrils: the D-periodicity and the groove depth were 65.7 +/- 0.8 nm and 1.46 +/- 0.50 nm in the corneal fibrils, and 67.3 +/- 1.1 nm and 6.16 +/- 1. 23 nm in the scleral fibrils.Conclusions: Surface topographic images of human corneal and scleral collagen fibrils were clearly obtained with the AFM. This technique provides quantitative information on the surface morphology of the collagen fibrils at high resolution.

Journal Article↗
PubMed1992

Atomic force microscopy of single- and double-stranded DNA.

A method has been developed for imaging single-stranded DNA with the atomic force microscope (AFM). phi X174 single-stranded DNA in formaldehyde on mica can be imaged in the AFM under propanol or butanol or in air. Measured lengths of most molecules are on the order of 1 mu, although occasionally more extended molecules with lengths of 1.7 to 1.9 mu are seen. Single-stranded DNA in the AFM generally appears lumpier than double-stranded DNA, even when extended. Images of double-stranded lambda DNA in the AFM show more sharp kinks and bends than are typically observed in the electron microscope. Dense, aggregated fields of double-stranded plasmids can be converted by gentle rinsing with hot water to well spread fields.

Bacteriophage lambda↗
PubMed1993

Structure of the extracellular surface of the gap junction by atomic force microscopy.

The extracellular surface of the gap junction cell-to-cell channels was imaged in phosphate-buffered saline with an atomic force microscope. The fully hydrated isolated gap junction membranes adsorbed to mica were irregular sheets approximately 1-2 microns across and 13.2 (+/- 1.3) nm thick. The top bilayer of the gap junction was dissected by increasing the force applied to the tip or sometimes by increasing the scan rate at moderate forces. The exposed extracellular surface revealed a hexagonal array with a center-to-center spacing of 9.4 (+/- 0.9) nm between individual channels (connexons). Images of individual connexons with a lateral resolution of < 3.5 nm, and in the best case approximately 2.5 nm, were reliably and reproducibly obtained with high-quality tips. These membrane channels protruded 1.4 (+/- 0.4) nm from the extracellular surface of the lipid membrane, and the atomic force microscope tip reached up to 0.7 nm into the pore, which opened up to a diameter of 3.8 (+/- 0.6) nm on the extracellular side.

Animals↗
PubMed2000

Amplitude curves and operating regimes in dynamic atomic force microscopy

The experimental dependence of the amplitude on the average tip-sample distance has been studied to understand the operation of an atomic force microscope with an amplitude modulation feedback. The amplitude curves can be classified in three major groups according to the existence or not of a local maximum and how the maximum is reached (steplike discontinuities vs. smooth transitions). A model describing the cantilever motion as a forced nonlinear oscillator allows to associate the features observed in the amplitude curves with the tip-sample interaction force. The model also allows to define two elemental tip-sample interaction regimes, attractive and repulsive. The presence of a local maximum in the amplitude curves is related to a transition between the attractive and the repulsive regime.

Journal Article↗
PubMed1994

Imaging human erythrocyte spectrin with atomic force microscopy.

Isolated spectrin covalently attached to a surface in a liquid environment as well as dried on mica has been studied with a contact-mode atomic force microscope. Both pyramidal and conical-type cantilever tip facets were used in the AFM. Our images show structures and give dimensions that correlate well with previous structural studies using transmission electron microscopy.

Erythrocyte Membrane↗
PubMed2002

Surface modification of an organosilane self-assembled monolayer on silicon substrates using atomic force microscopy: scanning probe electrochemistry toward nanolithography.

Organosilane self-assembled monolayers (SAMs) have been applied to resist materials for nanolithography based on scanning probe microscopy. An organosilane SAM was prepared on Si substrates from a precursor, that is octadecyltrimethoxysilane. Using an atomic force microscope with a conductive probe, current was injected from the probe into the SAM-covered Si substrate so that the SAM was locally degraded at the probe-contacting point. Nanoscale patterns drawn on the SAM was clearly imaged by lateral force microscopy. The patterning could be conducted in air while, in vacuum at the order of 10(-6) Torr, no detectable patterns were fabricated. The presence of adsorbed water at the probe/sample junction was confirmed to be crucial for the patterning of the SAM/Si. Its mechanism was, thus, ascribed to electrochemical reactions of both the SAM and Si with adsorbed water.

Journal Article↗
PubMed2000

Atomic force microscopy captures quantized plastic deformation in gold nanowires.

Scanning probe microscopy has become a powerful tool to detect structural changes in small clusters of atoms. Herein, we use an atomic force microscope to measure the length of gold nanowire structures during extension and compression cycles. We have found that nanowires elongate under force in quantized steps of up to three integer multiples of 1.76 A and that they shorten spontaneously in steps of 1.52 A. Our results can be explained by the sliding of crystal planes within the gold nanowires creating stacking faults that change the local structure from face-centered cubic to hexagonal close packed. Our data also show that there can be up to three simultaneous slip events, in good agreement with the tetrahedral arrangement of slip planes in a gold crystal. These experiments provide direct evidence for the mechanism underlying the plastic deformation of a nanowire. A similar approach can be used to examine the atomic events underlying the plastic failure of other metals and their alloys.

Journal Article↗
PubMed2001

Atomic force microscopy observations of in situ deformed materials: application to single crystals and thin films on substrates.

An experimental apparatus which consists of a compression machine interfaced with an atomic force microscope has been realized and allows the in situ observation of a sample surface under compressive stress. Taking advantage of the high resolution offered by this microscopy, the equipment is particularly suited both to analysing the fine slip line structure of deformed single crystals, providing interesting complementary information about plastic mechanisms taking place in the bulk, and to characterizing the mechanical behaviour of thin films on substrates with the investigation of the buckling phenomenon.

Journal Article↗
PubMed2001

Fabrication and characterization of metal-molecule-metal junctions by conducting probe atomic force microscopy.

Metal-molecule-metal junctions were fabricated by contacting Au-supported alkyl or benzyl thiol self-assembled monolayers (SAMs) with an Au-coated atomic force microscope (AFM) tip. The tip-SAM microcontact is approximately 15 nm(2), meaning the junction contains approximately 75 molecules. Current-voltage (I-V) characteristics of these junctions were probed as a function of SAM thickness and load applied to the microcontact. The measurements showed: (1) the I-V traces were linear over +/-0.3 V, (2) the junction resistance increased exponentially with alkyl chain length, (3) the junction resistance decreased with increasing load and showed two distinct power law scaling regimes, (4) resistances were a factor of 10 lower for junctions based on benzyl thiol SAMs compared to hexyl thiol SAMs having the same thickness, and (5) the junctions sustained fields up to 2 x 10(7) V/cm before breakdown. I-V characteristics determined for bilayer junctions involving alkane thiol-coated tips in contact with alkane thiol SAMs on Au also showed linear I-Vs over +/-0.3 V and the same exponential dependence on thickness. The I-V behavior and the exponential dependence of resistance on alkyl chain length are consistent with coherent, nonresonant electron tunneling across the SAM. The calculated conductance decay constant (beta) is 1.2 per methylene unit ( approximately 1.1 A(-)(1)) for both monolayer and bilayer junctions, in keeping with previous scanning tunneling microscope and electrochemical measurements of electron transfer through SAMs. These measurements show that conducting probe-AFM is a reliable method for fundamental studies of electron transfer through small numbers of molecules. The ability to vary the load on the microcontact is a unique characteristic of these junctions and opens opportunities for exploring electron transfer as a function of molecular deformation.

Journal Article↗
PubMed1992

Actin filament dynamics in living glial cells imaged by atomic force microscopy.

Observation of filamentous actin (F-actin) in living cells is currently limited to the resolution of the light microscope. Higher resolution procedures require sample fixation and preclude dynamic studies. The atomic force microscope (AFM) can image and manipulate samples at very high, sometimes atomic resolution by scanning a fine tip over the surface of interest and detecting physical interactions between the tip and sample. This study demonstrates that F-actin can be readily resolved in living cells with the AFM and that the dynamic properties of F-actin are easily observed.

Actin Cytoskeleton↗
PubMed2000

Atomic force microscopy of the three-dimensional crystal of membrane protein, OmpC porin.

Three-dimensional microcrystals of OmpC osmoporin were air-dried slowly and imaged in air with an atomic force microscope (AFM). The overall structural features in AFM images are in good agreement with the X-ray diffraction data of these OmpC osmoporin crystals: monoclinic P2(1) with the unit cell constants a=117.6 Å, b=110 Å, c=298.4 Å, beta=97 degrees. Such a good correspondence between X-ray diffraction and AFM data suggests that the slow and mild air-drying of these crystals did not induce any significant alterations in the crystal lattices as expected upon crystal dehydration. At the (100) crystal face, individual trimeric protein-detergent complexes were resolved. These results show the potential for studying the molecular structure of microcrystals of integral membrane proteins. This study also suggests that the crystal grew in a fashion of rapid two-dimensional expansion along the bc plane followed by a slow deposition along the a axis, perhaps as a rate-limiting nucleation process. Thus, AFM imaging of air-dried crystals would also be of considerable use in the early stages of a project to grow large three-dimensional crystals of membrane proteins suitable for high-resolution X-ray diffraction studies.

Journal Article↗
PubMed2004

Single-molecule recognition imaging microscopy.

Atomic force microscopy is a powerful and widely used imaging technique that can visualize single molecules and follow processes at the single-molecule level both in air and in solution. For maximum usefulness in biological applications, atomic force microscopy needs to be able to identify specific types of molecules in an image, much as fluorescent tags do for optical microscopy. The results presented here demonstrate that the highly specific antibody-antigen interaction can be used to generate single-molecule maps of specific types of molecules in a compositionally complex sample while simultaneously carrying out high-resolution topographic imaging. Because it can identify specific components, the technique can be used to map composition over an image and to detect compositional changes occurring during a process.

Animals↗
PubMed1992

Investigation of dialysis membranes with atomic force microscopy.

AFM was used to investigate dialysis membranes made of regenerated cellulose by the cuoxam process. The membranes were either Cuprophan or experimental samples, modified with different amounts of diethylaminoethylcellulose (DEAE). Atomic force microscopes with optical-lever detection systems were used to image the dry membranes in air as received from the manufacturer as well as wet membranes in a swollen state under water. Differences could be observed between modified and unmodified as well as between dry and wet membranes.

Cellulose↗
PubMed2006

Electrochemistry of conductive polymers 38. Electrodeposited poly(3,4-ethylenedioxy-thiophene) studied by current sensing atomic force microscopy.

Electrical and morphological properties of poly(3,4-ethylenedioxythiophene) (PEDOT) thin films electrodeposited on gold-on-silicon electrodes by galvanostatic, potentiostatic, and potentiodynamic methods have been determined using current sensing atomic force microscopic experiments. Surface morphologies and vertical conductivities of PEDOT films were affected by the experimental parameters including the preparation method, the current density, the potential, and the potentiodynamic cyclic number. Band gaps obtained from current-voltage curves of dedoped PEDOT were in excellent agreement with those obtained from absorption spectra. When the film thickness was increased on the gold-on-silicon electrode, the topographic images were not very well defined due to the high roughness but conductivities increased significantly in all the galvanostatically, potetiostatically, and potentiodynamically grown PEDOT films.

Journal Article↗
PubMed2005

Ferrocenylundecanethiol self-assembled monolayer charging correlates with negative differential resistance measured by conducting probe atomic force microscopy.

Electrical and mechanical properties of metal-molecule-metal junctions formed between Au-supported self-assembled monolayers (SAMs) of electroactive 11-ferrocenylundecanethiol (FcC(11)SH) and a Pt-coated atomic force microscope (AFM) tip have been measured using a conducting probe (CP) AFM in insulating alkane solution. Simultaneous and independent measurements of currents and bias-dependent adhesion forces under different applied tip biases between the conductive AFM probe and the FcC(11)SH SAMs revealed reversible peak-shaped current-voltage (I-V) characteristics and correlated maxima in the potential-dependent adhesion force. Trapped positive charges in the molecular junction correlate with high conduction in a feature showing negative differential resistance. Similar measurements on an electropassive 1-octanethiol SAM did not show any peaks in either adhesion force or I-V curves. A mechanism involving two-step resonant hole transfer through the occupied molecular orbitals (MOs) of ferrocene end groups via sequential oxidation and subsequent reduction, where a hole is trapped by the phonon relaxation, is proposed to explain the observed current-force correlation. These results suggest a new approach to probe charge-transfer involving electroactive groups on the nanoscale by measuring the adhesion forces as a function of applied bias in an electrolyte-free environment.

Journal Article↗