Contact Mode Atomic Force Microscopy Research Articles

Force Curve Measurement of Self-assembled Organic Monolayers Bound Covalently on Silicon (111)

Self-assembled monolayers of alkyl chains were prepared on silicon (111) surfaces. The surface was characterized by atomic force microscopy (AFM). Atomically flat terraces were observed in topographic images of the films with contact mode AFM, indicating the formation of highly ordered monolayers. Force curve measurements showed that the adhesion force between organic films and gold coated cantilever was much stronger comparing to the force on hydrogen terminated surfaces.

Investigating live and fixed epithelial and fibroblast cells by atomic force microscopy

Purpose. To investigate the use of atomic force microscopy (AFM) to image live and fixed cell in culture. Rabbit corneal fibroblasts, Chang conjunctival cells, and transformed human corneal epithelial cells were chosen so that AFM parameters could be set for future use in toxicologic and pharmacologic studies of ocular cells. Methods. Contact mode AFM was performed under air and in balanced salt solution (BSS) using live and fixed cells. All cell lines were imaged in the height mode for optimal resolution of cellular features. Results. Images of fixed cells showed no discernible differences in surface features when visualized in air or under physiologic solution. Structural differences were observed, however, between fixed and live cells in BSS. Although the AFM technique provides high quality images of live cells under BSS, sub-membrane features of live cells are more well-defined compared to fixed cells. It was also possible to image live cells in air if imaging was completed within 10 minutes of removal of the cells from culture medium. Images of cytoskeletal features under air were similar to those obtained under BSS. Conclusions. The atomic force microscopy technique can be used to study cells and provide sub-cellular details at resolution equal to or in some situations better than the scanning electron microscopy technique. However, parameters for imaging have to be tailored for individual experimental goals.

AFM and TEM Studies of Pt Nanoparticle Arrays Supported on Alumina Model Catalyst Prepared by Electron Beam Lithography

Pt nanoparticle arrays were fabricated by electron beam lithography as a model for supported catalysts. The adhesion strength of 28 nm Pt nanoparticles deposited on alumina has been studied using contact mode atomic force microscopy. On as-prepared samples, the metal nanoparticles were removed by the AFM tip with a force of approximately 30 nN. After heat treatment at 500°C in a vacuum, Pt nanoparticles could not be removed by the AFM tip, even at 4000 nN. The increase of adhesion upon heat treatment indicates stronger bonding between Pt and the support. TEM examination showed that the Pt nanoparticles were polycrystalline before any treatment, with the crystalline domain increasing significantly after heat treatment.

Mapping of Lateral Vibration Amplitude of the Tip in Contact Mode Atomic Force Microscopy

Mapping of Lateral Vibration Amplitude of the Tip in Contact Mode Atomic Force Microscopy

Nano-oxidation of titanium films with large atomically flat surfaces by means of voltage-modulated scanning probe microscopy

Titanium films with large atomically flat surfaces were produced by adapting the template stripping method. The morphology of these surfaces was characterized by contact mode atomic force microscopy and a root-mean-square roughness of 0.23 nm at a 1 µm2 area was obtained. Sample bias voltage modulation was used for nanometer-scale anodic oxidation of the Ti film. The oxide structures produced showed high reproducibility, with aspect ratios of 0.18 and 0.07 for voltage modulation and static pulse oxidation, respectively. The maximum height of oxide structures was found at ∼20 Hz voltage modulation frequency. Using a negative voltage on the sample in a combination with a Pt/Ir-coated tip enhanced further the aspect ratio and increased the oxide structure height. The influence of control parameters on the aspect ratio of the oxide structures was studied. Copyright © 2002 John Wiley & Sons, Ltd.

Effect of thiourea and saccharin on the roughness of electrodeposited ultrathin nickel and cobalt layers

Ni and Co films were produced by electrodeposition from plating baths containing either thiourea or saccharin. The effect of the organic additives on surface roughness was studied for the Ni-thiourea, Ni-saccharin, Co-thiourea and Co-saccharin systems. Layer thicknesses were varied from 1 to 10 nm and the additive concentration was varied from about 1 μM to 1 mM. Contact mode atomic force microscopy was used to measure both the root mean square (RMS) peak height (nm) and the areal peak density (μm−2) of each film. Although the RMS peak height and areal density were both influenced by film thickness and by additive concentration, the two roughness measures provide complementary information on the layer morphology and growth mechanisms. The four systems studied responded differently to changes in the concentration of the added organic. For example, for films 8 nm thick, addition of 1.22 × 10−3 thiourea reduced the Ni peak density about 80%; in contrast, the Co peak density was increased over 180% by addition of 1.25 × 10−3 thiourea. Use of peak height and peak density data to infer growth mechanisms for ultrathin films is discussed.

Preparation and Nanopatterning of Organic Monolayers Anchored Covalently to Silicon Surfaces.

Organic monolayers anchored to silicon surfaces via silicon-carbon covalent bonds were prepared by wet process and 2D-patterning of the surface was done by local oxidation with an atomic force microscope (AFM). The surfaces were anodized with a contact-mode AFM by applying a positive bias voltage to the surface with respect to the platinum-coated cantilever under ambient conditions, which resulted in nanometer-scale oxidation of the surfaces. The anodized areas were etched and terminated with hydrogen atoms by NH4F solution, in which various molecules having C=C bonds could be immobilized. We put allylamine molecules to which organic dyes such as fluorescein and porphyrin were anchored by chemical reactions. The intensity of luminescence varied depending on dopant concentration of the substrates possibly due to energy transfer between the dyes and substrates. The method demonstrated here is one of the promising way to fabricate 3 D-assemblies of molecular scale electronic devices on silicon with a stable interface.

Wet-Process Molecular Planting in A Specific Site on Silicon with Si-C Covalent Bonds

ABSTRACTAlkyl monolayers anchored covalently on silicon were prepared through the reaction between 1-alkenes and hydrogen-terminated silicon (111) surfaces. The 2D-patterning of the surface was done by local oxidation with an atomic force microscope (AFM) and 3D-molecular assemblies were fabricated by immobilization of molecules in a specific site of the patterned surface. The surfaces were anodized with a contact-mode AFM by applying a positive bias voltage to the surface with respect to the platinum-coated cantilever under ambient conditions, which resulted in nanometer-scale oxidation of surfaces. The anodized areas were etched and terminated with hydrogen atoms by NH4F solution, in which we could immobilize various molecules having C=C bonds. We put arylamine molecules to which organic dyes such as fluoroscein were anchored. The intensity of luminescence varied depending on dopant concentration of substrates. Luminescence was very weak on highly-doped silicon possibly due to effective energy transfer from dyes to substrates. The method demonstrated is one of the promising ways to fabricate 3D-assemblies of molecular-scale electronic devices with a stable interface on silicon.

Comparison of atomic force microscopy imaging methods and roughness determinations for a highly polished quartz surface

The morphology and microstructure of a polished quartz surface have been studied using atomic force microscopy (AFM) under ambient conditions, high vacuum and ultrahigh vacuum. The general features observed in air and vacuum environments are consistent, showing the existence of long corrugations, scratches, and holes. The root-mean-square surface roughness ranged from 0.2 to 0.4 nm. Imaging of hydrophilic surfaces, such as quartz, with contact-mode AFM can be problematic because of the presence of adsorbed water and contamination layers on the surface. Using tapping-mode AFM in air and noncontact mode AFM under vacuum, we are able to study the true structure of the surface; detection of more local features with higher resolution was achieved under ultrahigh-vacuum conditions.

Simulation of the observability of atomic defects by atomic force microscopy in contact and non-contact modes

Atomic force microscopy (AFM) scans of a crystal surface containing an atomic defect were simulated in both contact and non-contact regimes. When scanning in contact mode near a defect, the tip–sample force interaction experiences bifurcation of the lines of constant force. When the load force is small, the bifurcation causes the tip to be “pushed” out of the defect. However, if scan force is higher than some critical value (dependent upon the composition of the tip and sample) the AFM tip becomes “trapped” in the vicinity of defect. The trapped tip remains at the level of the vacancy and consequently crashes into the sample, as the scan continues. This results in either the tip apex being destroyed, or disruption of the crystal lattice around the defect. Both effects result in the “disappearance” of the defect from the scan images. The trap is intrinsic and cannot be avoided. For the case of non-contact mode, the tip position is driven by the scan force gradient rather than the force. Simulations show that for this case the trap does not exist and atomic defects will not be destroyed. This explains why atomic defects are generally not observed when using contact mode AFM, but are observed in non-contact AFM.

Tapping and contact mode imaging of native chromosomes and extraction of genomic DNA using AFM tips

It is very important both in medicine and biology to clarify the chromosomal structure to understand its functions. In a standard cytogenetic procedure, chromosomes are often fixed in a mixture of acetic acid and methanol. This process most likely changes the mechanical property of chromosomes. We adopted a method to prepare native and unfixed chromosomes from mouse 3T3 cells and used tapping and contact mode atomic force microscopy (AFM) to image and manipulate them. Modified AFM tips were used to image chromosomes in contact mode in air, and then the chromosome samples were immobilized on a substrate and placed in a buffer solution to pull out DNA–histone complexes from them after they were optimally treated with trypsin. From the AFM images, we could see several bands and granular structures on chromosomes. We obtained force curves indicating long fiber extensions from native chromosomes both with low (in high concentration of NaCl) and high forces (physiological conditions). The result suggested that the degree of chromosome condensation decreased in high concentration of salt. It agrees with the known fact of histone H1 dissociation in a high concentration of salt. We intend to pull out DNA–histone complexes from chromosomes for later molecular operations on them using an AFM.

Deformation of Lactococcus lactis surface in atomic force microscopy study

The surface of the bacterium Lactococcus lactis was investigated under water by contact mode atomic force microscopy (AFM) while varying the imaging parameters (imaging force, scanning velocity, scanning direction). Height images were three-dimensionally (3-D) reconstructed using GOCAD© software; this revealed grooves oriented along the scanning direction. Although the grooves were created by the scanning probe, they were not due to a single passage of the probe; the periodicity of the grooves was indeed always three to four times larger than the scanning line periodicity (ratio between the image size and the number of scanning lines). Upon repeated imaging at low force, the grooves were re-created each time with the same morphology; groove depth was increased at higher imaging force. The groove formation is tentatively explained by a deformation of the surface, due to compression or to adhesion, and its slow relaxation. While the grooves reveal a perturbation of the cell surface by the AFM probe, they provide pertinent information about the nanomechanical properties of the cell surface.

Early detection of cytotoxic events between hepatic natural killer cells and colon carcinoma cells as probed with the atomic force microscope

The atomic force microscope (AFM) is a powerful tool to investigate surface and submembranous structures of living cells under physiological conditions at high resolution. These properties enabled us to study the interaction between live hepatic natural killer (NK) cells, also called pit cells, and colon carcinoma cells in vitro by AFM. In addition, the staining for filamentous actin and DNA was performed and served as a reference, because actin and nuclear observations at the light microscopic level during the cytotoxic interaction between these two cell types have been presented earlier. In this study, we collected evidence that conjugation of hepatic NK cells with CC531s colon carcinoma cells results in a decreased binding of CC531s cells to the substratum as probed with the AFM in contact mode as early as 10 min after cell contact ( n=11). To avoid the lateral forces and smearing artefacts of contact mode AFM, non-contact imaging was performed on hepatic NK/CC531s cell conjugates, resulting in identical observations ( n=3). In contrast, the first cytotoxic signs, as determined with the nuclear staining dye Hoechst 33342, could be observed 3 h after the start of the co-culture. This study illustrates that the AFM can be used to probe early cytotoxic effects of effector to target cell contact in nearby physiological conditions. Other routine cytotoxicity tests detect the first cytotoxic effects after 1.5–3 h co-incubation at the earliest.

The application of laser scanning confocal microscopy to tribological research

Since the introduction of the first commercial systems in the early 1980s laser scanning confocal microscopy (LSCM) has become an established technique in biological and medical fields of research. To date the application of LSCM to metallurgical and tribological fields of research has been extremely limited. However, largely as a result of recent rapid advances in computer processing power, the modern LSCM system has become a flexible research tool with a broad range of capabilities, which are well suited to metallurgical research. In this article, the application of LSCM to the study of worn surfaces is discussed. Illustrations are presented which show how post-processing of confocal image stacks can be used to achieve a greatly extended depth of field thus enabling clear images of rough tribological surfaces to be constructed. Furthermore, illustrations of the use of LSCM for the quantification of surface topography are also presented which demonstrate that surface height profiles which faithfully reproduce the geometry of the real surface can be measured with a high degree of accuracy. The accuracy of profile surface roughness measurements via the LSCM has also been systematically investigated and compared with the results obtained from profiles measured using contact mode atomic force microscopy (AFM). The results presented show that, after suitable data post-processing to correct for tilt and extraneous signal noise, the results obtained using both techniques are in good agreement. Finally, the results of a study of rolling sliding contact wear surfaces of two high chromium content white cast irons are presented. The combination of extended focal depth imaging and topographical quantification afforded by the LSCM has proven capable of firmly establishing the wear mechanisms operating in these materials.

Polymer sandwiches: polyaniline films deposited on thiol-coated gold by chemical in situ method

We present a study of polyaniline chemical deposition in situ on gold electrodes coated with thiol monolayers containing different terminal groups: methyl, hydroxy, and carboxy. The differences in the deposition rates of polymers on various thiol monolayers were investigated with the quartz crystal microbalance (QCM) and UV-vis spectroscopy. It was found that the polymer is formed in the bulk of the polymerization solution and subsequently precipitated on the electrode surface. The synthesized polymers were studied by cyclic voltammetry. A novel method allowing for the investigation of the inner (thiol) layer integrity was proposed. It is based on the electrochemical oxidation of the gold substrate and formation of complexes with chloride ions incorporated in the polymer films. The observation of a characteristic AuCl 4 − reduction peak allows for the determination whether the polymer is adjacent just to the surface of gold or it is separated from the bare metal by thiol monolayer. It was found that the deposition of polymer on the thiol monolayers with terminal methyl and carboxy groups leads to the formation of sandwich-like systems with the thiol monolayer interposed between the polymer layer and the gold surface, while the chemical polymerization on hydroxy-ended thiols induces partial desorption of the monolayer; thus, the polymer is directly adjacent to the bare gold over large areas of the electrode surface. The morphology of the synthesized polymer was also investigated by contact mode atomic force microscopy (AFM).

The interplay between surface micro-topography and -mechanics of type I collagen fibrils in air and aqueous media: An atomic force microscopy study

Calf skin type I collagen fibrils were regenerated from acidic solution and imaged with contact mode atomic force microscopy in air, water, and buffer solution. When imaged in air at a contact force of 20-150 nN, collagen fibrils exhibited a distinct transverse banding pattern with a period of 65 nm, consisting of high ridges and shallow grooves. The force dependence of the images suggests that such banding pattern is attributed to the transverse contraction of the fibril upon dehydration during sample preparation, which reflects the tangential mass density across the fibril. Imaging in water and phosphate buffer solution at a contact force of 15-80 nN revealed hydrated collagen fibrils with smooth surfaces. The rigidity of the collagen fibrils decreased considerably upon hydration. Scanning the cantilever tip in an aqueous medium at a contact force of 90-280 nN enabled us to probe subunit arrangement in the bulk region of the collagen fibril. The results indicate that the molecular assembly in the hydrated fibril is akin to that in the intact form. The image resolution was improved by stabilizing the collagen molecules through crosslinking with glutaraldehyde, which served to resolve microfibril-like structure on the fibril surface.

NANOSTRUCTURE OF THE DIATOM FRUSTULE AS REVEALED BY ATOMIC FORCE AND SCANNING ELECTRON MICROSCOPY

The cell wall (frustule) of the freshwater diatom Pinnularia viridis (Nitzsch) Ehrenberg is composed of an assembly of highly silicified components and associated organic layers. We used atomic force microscopy (AFM) to investigate the nanostructure and relationship between the outermost surface organics and the siliceous frustule components of live diatoms under natural hydrated conditions. Contact mode AFM imaging revealed that the walls were coated in a thick mucilaginous material that was interrupted only in the vicinity of the raphe fissure. Analysis of this mucilage by force mode AFM demonstrated it to be a nonadhesive, soft, and compressible material. Application of greater force to the sample during repeated scanning enabled the mucilage to be swept from the hard underlying siliceous components and piled into columns on either side of the scan area by the scanning action of the tip. The mucilage columns remained intact for several hours without dissolving or settling back onto the cleaned valve surface, thereby revealing a cohesiveness that suggested a degree of cross‐linking. The hard silicified surfaces of the diatom frustule appeared to be relatively smooth when living cells were imaged by AFM or when field‐emission SEM was used to image chemically cleaned walls. AFM analysis of P. viridis frustules cleaved in cross‐section revealed the nanostructure of the valve silica to be composed of a conglomerate of packed silica spheres that were 44.8 ± 0.7 nm in diameter. The silica spheres that comprised the girdle band biosilica were 40.3 ± 0.8 nm in diameter. Analysis of another heavily silicified diatom, Hantzschia amphioxys (Ehrenberg) Grunow, showed that the valve biosilica was composed of packed silica spheres that were 37.1 ± 1.4 nm and that silica particles from the girdle bands were 38.1 ± 0.5 nm. These results showed little variation in the size range of the silica particles within a particular frustule component (valve or girdle band), but there may be differences in particle size between these components within a diatom frustule and significant differences are found between species.

Atomic force microscopy study of salivary pellicles formed on enamel and glass in vivo

This study was performed to evaluate the use of atomic force microscopy (AFM) in examining the surface of the adsorbed layer of salivary proteins (salivary pellicle) formed in vivo on dental enamel and glass surfaces. Enamel and glass test pieces were attached to the buccal surfaces of the upper first molar teeth in two adults using removable intraoral splints. The splints were carried intraorally over periods ranging from 10 min to1 h. Using the contact mode of AFM, pellicle structures could be recognised on intraorally exposed specimens compared to nonexposed enamel and glass surfaces. The surface of the adsorbed salivary pellicle was characterised by a dense globular appearance. The diameter of the globulelike protein aggregates adsorbed onto enamel and glass varied between 80 and 200 nm and 80 and 150 nm, respectively. The structure of the adsorbed protein layer was clearly visible on glass surfaces, even though minor differences in the protein layer between glass and enamel specimens were observed. This study indicates that AFM is a powerful tool for high-resolution examination of the salivary pellicle surface structure in its native (hydrated) state. AFM avoids artefacts due to fixing, dehydration and sputter-coating which occur with scanning electron microscopic analyses.

Modulating artificial membrane morphology: pH-induced chromatic transition and nanostructural transformation of a bolaamphiphilic conjugated polymer from blue helical ribbons to red nanofibers.

Design and characterization of helical ribbon assemblies of a bolaamphiphilic conjugated polymer and their color-coded transformation into nanofibers are described. An L-glutamic acid modified bolaamphiphilic diacetylene lipid was synthesized and self-assembled into right-handed helical ribbons with micron scale length and nano scale thickness under mild conditions. The ribbon structures were further stabilized by polymerizing well-aligned diacetylene units to form bisfunctional polydiacetylenes (PDAs). Transitions from flat sheets to helical ribbons and tubes were observed by transmission electron microscopy. The helical ribbons appear to originate from the rupture of flat sheets along domain edges and the peeling off between stacked lipid layers. These results point to the applicability of chiral packing theory in bolaamphiphilic supramolecular assemblies. Contact mode atomic force microscopy observations revealed that high order existed in the surface packing arrangement. Hexagonal and pseudorectangular packings were observed in flat and twisted regions of the ribbons, respectively, suggesting a correlation between microscopic morphologies and nanoscopic packing arrangements. The tricarboxylate functionalities of the bolaamphiphilic lipid provide a handle for the manipulation of the bisfunctional PDAs' morphology. Increasing solution pH caused the fraying of helical ribbons into nanofibers accompanied by a sharp blue-to-red chromatic transition. A dramatic change in circular dichroism spectra was observed during this process, suggesting the loss of chirality in packing. A model is proposed to account for the pH-induced morphological change and chromatic transition. The color-coded transition between two distinct microstructures would be useful in the design of sensors and other "smart" nanomaterials requiring defined molecular templates.

Use of in-situ atomic force microscopy to monitor the biodegradation of polyhydroxyalkanoates (PHAs)

Thin films of a co-polymer mixture of poly 3-hydroxybutyrate and poly 3-hydroxyvalerate P(3HB-3HV) were spun-cast onto glass slides resulting in 35 nm thick layers with a spherulitic microstructure. An untyped strain of Streptomyces sp. bacteria was isolated from soil samples, and it's PHA depolymerase was used to degrade the P(3HB-3HV) thin films. Both ex-situ and in-situ atomic force microscopy (AFM) biodegradation studies were performed to determine the kinetics of the biodegradation over the course of three hours at room temperature. Ex-situ AFM was performed in Tapping Mode and in-situ AFM was performed in the PHA depolymerase using contact mode AFM in the liquid cell, allowing for the real-time analysis of P(3HB-3HV) biodegradation. Biodegradation is observed uniformly throughout the surface, and can be observed within 30 min. of depolymerase exposure. In-situ AFM analysis yields a linear degradation rate as a function of time, while the ex-situ study suggests a more complex kinetics.