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1.  Effect of the tip state during qPlus noncontact atomic force microscopy of Si(100) at 5 K: Probing the probe 
Background: Noncontact atomic force microscopy (NC-AFM) now regularly produces atomic-resolution images on a wide range of surfaces, and has demonstrated the capability for atomic manipulation solely using chemical forces. Nonetheless, the role of the tip apex in both imaging and manipulation remains poorly understood and is an active area of research both experimentally and theoretically. Recent work employing specially functionalised tips has provided additional impetus to elucidating the role of the tip apex in the observed contrast.
Results: We present an analysis of the influence of the tip apex during imaging of the Si(100) substrate in ultra-high vacuum (UHV) at 5 K using a qPlus sensor for noncontact atomic force microscopy (NC-AFM). Data demonstrating stable imaging with a range of tip apexes, each with a characteristic imaging signature, have been acquired. By imaging at close to zero applied bias we eliminate the influence of tunnel current on the force between tip and surface, and also the tunnel-current-induced excitation of silicon dimers, which is a key issue in scanning probe studies of Si(100).
Conclusion: A wide range of novel imaging mechanisms are demonstrated on the Si(100) surface, which can only be explained by variations in the precise structural configuration at the apex of the tip. Such images provide a valuable resource for theoreticians working on the development of realistic tip structures for NC-AFM simulations. Force spectroscopy measurements show that the tip termination critically affects both the short-range force and dissipated energy.
PMCID: PMC3304327  PMID: 22428093
force spectroscopy; image contrast; noncontact AFM; qPlus; Si(001); Si(100); tip (apex) structure
2.  Nanoscale Cell Wall Deformation Impacts Long-Range Bacterial Adhesion Forces on Surfaces 
Adhesion of bacteria occurs on virtually all natural and synthetic surfaces and is crucial for their survival. Once they are adhering, bacteria start growing and form a biofilm, in which they are protected against environmental attacks. Bacterial adhesion to surfaces is mediated by a combination of different short- and long-range forces. Here we present a new atomic force microscopy (AFM)-based method to derive long-range bacterial adhesion forces from the dependence of bacterial adhesion forces on the loading force, as applied during the use of AFM. The long-range adhesion forces of wild-type Staphylococcus aureus parent strains (0.5 and 0.8 nN) amounted to only one-third of these forces measured for their more deformable isogenic Δpbp4 mutants that were deficient in peptidoglycan cross-linking. The measured long-range Lifshitz-Van der Waals adhesion forces matched those calculated from published Hamaker constants, provided that a 40% ellipsoidal deformation of the bacterial cell wall was assumed for the Δpbp4 mutants. Direct imaging of adhering staphylococci using the AFM peak force-quantitative nanomechanical property mapping imaging mode confirmed a height reduction due to deformation in the Δpbp4 mutants of 100 to 200 nm. Across naturally occurring bacterial strains, long-range forces do not vary to the extent observed here for the Δpbp4 mutants. Importantly, however, extrapolating from the results of this study, it can be concluded that long-range bacterial adhesion forces are determined not only by the composition and structure of the bacterial cell surface but also by a hitherto neglected, small deformation of the bacterial cell wall, facilitating an increase in contact area and, therewith, in adhesion force.
PMCID: PMC3911093  PMID: 24212582
3.  Probing three-dimensional surface force fields with atomic resolution: Measurement strategies, limitations, and artifact reduction 
Noncontact atomic force microscopy (NC-AFM) is being increasingly used to measure the interaction force between an atomically sharp probe tip and surfaces of interest, as a function of the three spatial dimensions, with picometer and piconewton accuracy. Since the results of such measurements may be affected by piezo nonlinearities, thermal and electronic drift, tip asymmetries, and elastic deformation of the tip apex, these effects need to be considered during image interpretation.
In this paper, we analyze their impact on the acquired data, compare different methods to record atomic-resolution surface force fields, and determine the approaches that suffer the least from the associated artifacts. The related discussion underscores the idea that since force fields recorded by using NC-AFM always reflect the properties of both the sample and the probe tip, efforts to reduce unwanted effects of the tip on recorded data are indispensable for the extraction of detailed information about the atomic-scale properties of the surface.
PMCID: PMC3458610  PMID: 23019560
atomic force microscopy; force spectroscopy; NC-AFM; three-dimensional atomic force microscopy; tip asymmetry; tip elasticity
4.  Atomic Force Microscopy Application in Biological Research: A Review Study 
Atomic force microscopy (AFM) is a three-dimensional topographic technique with a high atomic resolution to measure surface roughness. AFM is a kind of scanning probe microscope, and its near-field technique is based on the interaction between a sharp tip and the atoms of the sample surface. There are several methods and many ways to modify the tip of the AFM to investigate surface properties, including measuring friction, adhesion forces and viscoelastic properties as well as determining the Young modulus and imaging magnetic or electrostatic properties. The AFM technique can analyze any kind of samples such as polymers, adsorbed molecules, films or fibers, and powders in the air whether in a controlled atmosphere or in a liquid medium. In the past decade, the AFM has emerged as a powerful tool to obtain the nanostructural details and biomechanical properties of biological samples, including biomolecules and cells. The AFM applications, techniques, and -in particular- its ability to measure forces, are not still familiar to most clinicians. This paper reviews the literature on the main principles of the AFM modality and highlights the advantages of this technique in biology, medicine, and- especially- dentistry. This literature review was performed through E-resources, including Science Direct, PubMed, Blackwell Synergy, Embase, Elsevier, and Scholar Google for the references published between 1985 and 2010.
PMCID: PMC3700051  PMID: 23825885
Atomic force microscopy; Scanning tunneling microscopy; Scanning probe microscopy; Dental; Biological
5.  The role of surface corrugation and tip oscillation in single-molecule manipulation with a non-contact atomic force microscope 
Scanning probe microscopy (SPM) plays an important role in the investigation of molecular adsorption. The possibility to probe the molecule–surface interaction while tuning its strength through SPM tip-induced single-molecule manipulation has particularly promising potential to yield new insights. We recently reported experiments, in which 3,4,9,10-perylene-tetracarboxylic-dianhydride (PTCDA) molecules were lifted with a qPlus-sensor and analyzed these experiments by using force-field simulations. Irrespective of the good agreement between the experiment and those simulations, systematic inconsistencies remained that we attribute to effects omitted from the initial model. Here we develop a more realistic simulation of single-molecule manipulation by non-contact AFM that includes the atomic surface corrugation, the tip elasticity, and the tip oscillation amplitude. In short, we simulate a full tip oscillation cycle at each step of the manipulation process and calculate the frequency shift by solving the equation of motion of the tip. The new model correctly reproduces previously unexplained key features of the experiment, and facilitates a better understanding of the mechanics of single-molecular junctions. Our simulations reveal that the surface corrugation adds a positive frequency shift to the measurement that generates an apparent repulsive force. Furthermore, we demonstrate that the scatter observed in the experimental data points is related to the sliding of the molecule across the surface.
PMCID: PMC3943512  PMID: 24605287
atomic force microscopy (AFM); force-field model; 3,4,9,10-perylene-tetracarboxylic-dianhydride (PTCDA); qPlus; single-molecule manipulation
6.  Statistical Analysis of Long- and Short-Range Forces Involved in Bacterial Adhesion to Substratum Surfaces as Measured Using Atomic Force Microscopy ▿  
Applied and Environmental Microbiology  2011;77(15):5065-5070.
Surface thermodynamic analyses of microbial adhesion using measured contact angles on solid substrata and microbial cell surfaces are widely employed to determine the nature of the adhesion forces, i.e., the interplay between Lifshitz-van der Waals and acid-base forces. While surface thermodynamic analyses are often viewed critically, atomic force microscopy (AFM) can also provide information on the nature of the adhesion forces by means of Poisson analysis of the measured forces. This review first presents a description of Poisson analysis and its underlying assumptions. The data available from the literature for different combinations of bacterial strains and substrata are then summarized, leading to the conclusion that bacterial adhesion to surfaces is generally dominated by short-range, attractive acid-base interactions, in combination with long-range, weaker Lifshitz-van der Waals forces. This is in line with the findings of surface thermodynamic analyses of bacterial adhesion. Comparison with single-molecule ligand-receptor forces from the literature suggests that the short-range-force contribution from Poisson analysis involves a discrete adhesive bacterial cell surface site rather than a single molecular force. The adhesion force arising from these cell surface sites and the number of sites available may differ from strain to strain. Force spectroscopy, however, involves the tedious task of identifying the minor peaks in the AFM retraction force-distance curve. This step can be avoided by carrying out Poisson analysis on the work of adhesion, which can also be derived from retraction force-distance curves. This newly proposed way of performing Poisson analysis confirms that multiple molecular bonds, rather than a single molecular bond, contribute to a discrete adhesive bacterial cell surface site.
PMCID: PMC3147456  PMID: 21642399
7.  Study of the Effect of Nanoparticles and Surface Morphology on Reverse Osmosis and Nanofiltration Membrane Productivity 
Membranes  2013;3(3):196-225.
To evaluate the significance of reverse osmosis (RO) and nanofiltration (NF) surface morphology on membrane performance, productivity experiments were conducted using flat-sheet membranes and three different nanoparticles, which included SiO2, TiO2 and CeO2. In this study, the productivity rate was markedly influenced by membrane surface morphology. Atomic force microscopy (AFM) analysis of membrane surfaces revealed that the higher productivity decline rates associated with polyamide RO membranes as compared to that of a cellulose acetate NF membrane was due to the inherent ridge-and-valley morphology of the active layer. The unique polyamide active layer morphology was directly related to the surface roughness, and was found to contribute to particle accumulation in the valleys causing a higher flux decline than in smoother membranes. Extended RO productivity experiments using laboratory grade water and diluted pretreated seawater were conducted to compare the effect that different nanoparticles had on membrane active layers. Membrane flux decline was not affected by particle type when the feed water was laboratory grade water. On the other hand, membrane productivity was affected by particle type when pretreated diluted seawater served as feed water. It was found that CeO2 addition resulted in the least observable flux decline, followed by SiO2 and TiO2. A productivity simulation was conducted by fitting the monitored flux data into a cake growth rate model, where the model was modified using a finite difference method to incorporate surface thickness variation into the analysis. The ratio of cake growth term (k1) and particle back diffusion term (k2) was compared in between different RO and NF membranes. Results indicated that k2 was less significant for surfaces that exhibited a higher roughness. It was concluded that the valley areas of thin-film membrane surfaces have the ability to capture particles, limiting particle back diffusion.
PMCID: PMC4021943  PMID: 24956946
productivity; nanoparticles; reverse osmosis; nanofiltration; cake growth model
8.  Surface Modifications by Field Induced Diffusion 
PLoS ONE  2012;7(1):e30106.
By applying a voltage pulse to a scanning tunneling microscope tip the surface under the tip will be modified. We have in this paper taken a closer look at the model of electric field induced surface diffusion of adatoms including the van der Waals force as a contribution in formations of a mound on a surface. The dipole moment of an adatom is the sum of the surface induced dipole moment (which is constant) and the dipole moment due to electric field polarisation which depends on the strength and polarity of the electric field. The electric field is analytically modelled by a point charge over an infinite conducting flat surface. From this we calculate the force that cause adatoms to migrate. The calculated force is small for voltage used, typical 1 pN, but due to thermal vibration adatoms are hopping on the surface and even a small net force can be significant in the drift of adatoms. In this way we obtain a novel formula for a polarity dependent threshold voltage for mound formation on the surface for positive tip. Knowing the voltage of the pulse we then can calculate the radius of the formed mound. A threshold electric field for mound formation of about 2 V/nm is calculated. In addition, we found that van der Waals force is of importance for shorter distances and its contribution to the radial force on the adatoms has to be considered for distances smaller than 1.5 nm for commonly used voltages.
PMCID: PMC3258261  PMID: 22253894
9.  A measurement of the hysteresis loop in force-spectroscopy curves using a tuning-fork atomic force microscope 
Measurements of the frequency shift versus distance in noncontact atomic force microscopy (NC-AFM) allow measurements of the force gradient between the oscillating tip and a surface (force-spectroscopy measurements). When nonconservative forces act between the tip apex and the surface the oscillation amplitude is damped. The dissipation is caused by bistabilities in the potential energy surface of the tip–sample system, and the process can be understood as a hysteresis of forces between approach and retraction of the tip. In this paper, we present the direct measurement of the whole hysteresis loop in force-spectroscopy curves at 77 K on the PTCDA/Ag/Si(111) √3 × √3 surface by means of a tuning-fork-based NC-AFM with an oscillation amplitude smaller than the distance range of the hysteresis loop. The hysteresis effect is caused by the making and breaking of a bond between PTCDA molecules on the surface and a PTCDA molecule at the tip. The corresponding energy loss was determined to be 0.57 eV by evaluation of the force–distance curves upon approach and retraction. Furthermore, a second dissipation process was identified through the damping of the oscillation while the molecule on the tip is in contact with the surface. This dissipation process occurs mainly during the retraction of the tip. It reaches a maximum value of about 0.22 eV/cycle.
PMCID: PMC3323909  PMID: 22496993
atomic force microscopy; energy dissipation; force spectroscopy; hysteresis loop; PTCDA/Ag/Si(111) √3 × √3
10.  Energy dissipation of nanoconfined hydration layer: Long-range hydration on the hydrophilic solid surface 
Scientific Reports  2014;4:6499.
The hydration water layer (HWL), a ubiquitous form of water on the hydrophilic surfaces, exhibits anomalous characteristics different from bulk water and plays an important role in interfacial interactions. Despite extensive studies on the mechanical properties of HWL, one still lacks holistic understanding of its energy dissipation, which is critical to characterization of viscoelastic materials as well as identification of nanoscale dissipation processes. Here we address energy dissipation of nanoconfined HWL between two atomically flat hydrophilic solid surfaces (area of ~120 nm2) by small amplitude-modulation, noncontact atomic force microscopy. Based on the viscoelastic hydration-force model, the average dissipation energy is ~1 eV at the tapping amplitude (~0.1 nm) of the tip. In particular, we determine the accurate HWL thickness of ~6 layers of water molecules, as similarly observed on biological surfaces. Such a long-range interaction of HWL should be considered in the nanoscale phenomena such as friction, collision and self-assembly.
PMCID: PMC4179125  PMID: 25267426
11.  Nanomechanical properties of α-synuclein amyloid fibrils: a comparative study by nanoindentation, harmonic force microscopy, and Peakforce QNM 
Nanoscale Research Letters  2011;6(1):270.
We report on the use of three different atomic force spectroscopy modalities to determine the nanomechanical properties of amyloid fibrils of the human α-synuclein protein. α-Synuclein forms fibrillar nanostructures of approximately 10 nm diameter and lengths ranging from 100 nm to several microns, which have been associated with Parkinson's disease. Atomic force microscopy (AFM) has been used to image the morphology of these protein fibrils deposited on a flat surface. For nanomechanical measurements, we used single-point nanoindentation, in which the AFM tip as the indenter is moved vertically to the fibril surface and back while the force is being recorded. We also used two recently developed AFM surface property mapping techniques: Harmonic force microscopy (HarmoniX) and Peakforce QNM. These modalities allow extraction of mechanical parameters of the surface with a lateral resolution and speed comparable to tapping-mode AFM imaging. Based on this phenomenological study, the elastic moduli of the α-synuclein fibrils determined using these three different modalities are within the range 1.3-2.1 GPa. We discuss the relative merits of these three methods for the determination of the elastic properties of protein fibrils, particularly considering the differences and difficulties of each method.
PMCID: PMC3211334  PMID: 21711775
12.  Probing softness of the parietal pleural surface at the micron scale 
Journal of biomechanics  2011;44(14):2558-2564.
The pleural surfaces of the chest wall and lung slide against each other, lubricated by pleural fluid. During sliding motion of soft tissues, shear induced hydrodynamic pressure deforms the surfaces, promoting uniformity of the fluid layer thickness, thereby reducing friction. To assess pleural deformability at length scales comparable to pleural fluid thickness, we measured the modulus of the parietal pleura of rat chest wall using atomic force microscopy (AFM) to indent the pleural surface with spheres (radius 2.5 µm and 5 µm). The pleura exhibited two distinct indentation responses depending on location, reflecting either homogeneous or significantly heterogeneous tissue properties. We found an elastic modulus of 0.38–0.95 kPa, lower than the values measured using flat-ended cylinders > 100 µm radii (Gouldstone et al., 2003, Journal of Applied Physiology 95, 2345–2349). Interestingly, the pleura exhibited a three-fold higher modulus when probed using 2.5 µm vs. 5 µm spherical tips at the same normalized depth, confirming depth dependent inhomogeneous elastic properties. The observed softness of the pleura supports the hypothesis that unevenness of the pleural surface on this scale is smoothed by local hydrodynamic pressure.
PMCID: PMC3168578  PMID: 21820660
Parietal pleura; AFM; Hertzian indentation; Elastohydrodynamic lubrication; Rat
13.  An NC-AFM and KPFM study of the adsorption of a triphenylene derivative on KBr(001) 
The adsorption on KBr(001) of a specially designed molecule, consisting of a flat aromatic triphenylene core equipped with six flexible propyl chains ending with polar cyano groups, is investigated by using atomic force microscopy in the noncontact mode (NC-AFM) coupled to Kelvin probe force microscopy (KPFM) in ultrahigh vacuum at room temperature. Two types of monolayers are identified, one in which the molecules lie flat on the surface (MLh) and another in which they stand approximately upright (MLv). The Kelvin voltage on these two structures is negatively shifted relative to that of the clean KBr surface, revealing the presence of surface dipoles with a component pointing along the normal to the surface. These findings are interpreted with the help of numerical simulations. It is shown that the surface–molecule interaction is dominated by the electrostatic interaction of the cyano groups with the K+ ions of the substrate. The molecule is strongly adsorbed in the MLh structure with an adsorption energy of 1.8 eV. In the MLv layer, the molecules form π-stacked rows aligned along the polar directions of the KBr surface. In these rows, the molecules are less strongly bound to the substrate, but the structure is stabilized by the strong intermolecular interaction due to π-stacking.
PMCID: PMC3323911  PMID: 22496995
atomic force microscopy; insulating surfaces; Kelvin force probe microscopy; molecular adsorption
14.  Uncertainties in forces extracted from non-contact atomic force microscopy measurements by fitting of long-range background forces 
In principle, non-contact atomic force microscopy (NC-AFM) now readily allows for the measurement of forces with sub-nanonewton precision on the atomic scale. In practice, however, the extraction of the often desired ‘short-range’ force from the experimental observable (frequency shift) is often far from trivial. In most cases there is a significant contribution to the total tip–sample force due to non-site-specific van der Waals and electrostatic forces. Typically, the contribution from these forces must be removed before the results of the experiment can be successfully interpreted, often by comparison to density functional theory calculations. In this paper we compare the ‘on-minus-off’ method for extracting site-specific forces to a commonly used extrapolation method modelling the long-range forces using a simple power law. By examining the behaviour of the fitting method in the case of two radically different interaction potentials we show that significant uncertainties in the final extracted forces may result from use of the extrapolation method.
PMCID: PMC3999863  PMID: 24778964
background subtraction; DFM; F(z); force; atomic resolution; NC-AFM; Si(111); STM; van der Waals
15.  Macroscale adhesion of gecko setae reflects nanoscale differences in subsurface composition 
Surface energies are commonly used to determine the adhesion forces between materials. However, the component of surface energy derived from long-range forces, such as van der Waals forces, depends on the material's structure below the outermost atomic layers. Previous theoretical results and indirect experimental evidence suggest that the van der Waals energies of subsurface layers will influence interfacial adhesion forces. We discovered that nanometre-scale differences in the oxide layer thickness of silicon wafers result in significant macroscale differences in the adhesion of isolated gecko setal arrays. Si/SiO2 bilayer materials exhibited stronger adhesion when the SiO2 layer is thin (approx. 2 nm). To further explore how layered materials influence adhesion, we functionalized similar substrates with an octadecyltrichlorosilane monolayer and again identified a significant influence of the SiO2 layer thickness on adhesion. Our theoretical calculations describe how variation in the SiO2 layer thickness produces differences in the van der Waals interaction potential, and these differences are reflected in the adhesion mechanics. Setal arrays used as tribological probes provide the first empirical evidence that the ‘subsurface energy’ of inhomogeneous materials influences the macroscopic surface forces.
PMCID: PMC3565786  PMID: 22993246
van der Waals forces; gecko adhesion; surface energy
16.  Tumor suppressor protein SMAR1 modulates the roughness of cell surface: combined AFM and SEM study 
BMC Cancer  2009;9:350.
Imaging tools such as scanning electron microscope (SEM) and atomic force microscope (AFM) can be used to produce high-resolution topographic images of biomedical specimens and hence are well suited for imaging alterations in cell morphology. We have studied the correlation of SMAR1 expression with cell surface smoothness in cell lines as well as in different grades of human breast cancer and mouse tumor sections.
We validated knockdown and overexpression of SMAR1 using RT-PCR as well as Western blotting in human embryonic kidney (HEK) 293, human breast cancer (MCF-7) and mouse melanoma (B16F1) cell lines. The samples were then processed for cell surface roughness studies using atomic force microscopy (AFM) and scanning electron microscopy (SEM). The same samples were used for microarray analysis as well. Tumors sections from control and SMAR1 treated mice as well as tissues sections from different grades of human breast cancer on poly L-lysine coated slides were used for AFM and SEM studies.
Tumor sections from mice injected with melanoma cells showed pronounced surface roughness. In contrast, tumor sections obtained from nude mice that were first injected with melanoma cells followed by repeated injections of SMAR1-P44 peptide, exhibited relatively smoother surface profile. Interestingly, human breast cancer tissue sections that showed reduced SMAR1 expression exhibited increased surface roughness compared to the adjacent normal breast tissue. Our AFM data establishes that treatment of cells with SMAR1-P44 results into increase in cytoskeletal volume that is supported by comparative gene expression data showing an increase in the expression of specific cytoskeletal proteins compared to the control cells. Altogether, these findings indicate that tumor suppressor function of SMAR1 might be exhibited through smoothening of cell surface by regulating expression of cell surface proteins.
Tumor suppressor protein SMAR1 might be used as a phenotypic differentiation marker between cancerous and non-cancerous cells.
PMCID: PMC2765988  PMID: 19799771
17.  Noncontact atomic force microscopy study of the spinel MgAl2O4(111) surface 
Based on high-resolution noncontact atomic force microscopy (NC-AFM) experiments we reveal a detailed structural model of the polar (111) surface of the insulating ternary metal oxide, MgAl2O4 (spinel). NC-AFM images reveal a 6√3×6√3R30° superstructure on the surface consisting of patches with the original oxygen-terminated MgAl2O4(111) surface interrupted by oxygen-deficient areas. These observations are in accordance with previous theoretical studies, which predict that the polarity of the surface can be compensated by removal of a certain fraction of oxygen atoms. However, instead of isolated O vacancies, it is observed that O is removed in a distinct pattern of line vacancies reflected by the underlying lattice structure. Consequently, by the creation of triangular patches in a 6√3×6√3R30° superstructure, the polar-stabilization requirements are met.
PMCID: PMC3323907  PMID: 22496991
aluminium oxide; metal oxide surfaces; noncontact atomic force microscopy (NC-AFM); polar surfaces; reconstructions; spinel
18.  High-resolution dynamic atomic force microscopy in liquids with different feedback architectures 
The recent achievement of atomic resolution with dynamic atomic force microscopy (dAFM) [Fukuma et al., Appl. Phys. Lett. 2005, 87, 034101], where quality factors of the oscillating probe are inherently low, challenges some accepted beliefs concerning sensitivity and resolution in dAFM imaging modes. Through analysis and experiment we study the performance metrics for high-resolution imaging with dAFM in liquid media with amplitude modulation (AM), frequency modulation (FM) and drive-amplitude modulation (DAM) imaging modes. We find that while the quality factors of dAFM probes may deviate by several orders of magnitude between vacuum and liquid media, their sensitivity to tip–sample forces can be remarkable similar. Furthermore, the reduction in noncontact forces and quality factors in liquids diminishes the role of feedback control in achieving high-resolution images. The theoretical findings are supported by atomic-resolution images of mica in water acquired with AM, FM and DAM under similar operating conditions.
PMCID: PMC3596120  PMID: 23503468
atomic force microscopy; dAFM; high-resolution; liquids
19.  Correlation between Desorption Force Measured by Atomic Force Microscopy and Adsorption Free Energy Measured by Surface Plasmon Resonance Spectroscopy for Peptide–Surface Interactions 
Surface Plasmon resonance (SPR) spectroscopy is a useful technique for thermodynamically characterizing peptide–surface interactions; however, its usefulness is limited to the types of surfaces that can readily be formed as thin layers in nanometer scale on metallic biosensor substrates. Atomic force microscopy (AFM), on the other hand, can be used with any microscopically flat surface, thus making it more versatile for studying peptide–surface interactions. AFM, however, has the drawback of data interpretation due to questions regarding peptide-to-probe–tip density. This problem could be overcome if results from a standardized AFM method could be correlated with SPR results for a similar set of peptide–surface interactions so that AFM studies using the standardized method could be extended to characterize peptide–surface interactions for surfaces that are not amenable for characterization by SPR. In this paper, we present the development and application of an AFM method to measure adsorption forces for host–guest peptides sequence on surfaces consisting of alkanethiol self–assembled monolayers (SAMs) with different functionality. The results from these studies show that a linear correlation exists between these data and the adsorption free energy (ΔG°ads) values associated with a similar set of peptide–surface systems available from SPR measurements. These methods will be extremely useful to thermodynamically characterize the adsorption behavior for peptides on a much broader range of surfaces than can be used with SPR to provide information related to understanding protein adsorption behavior to these surfaces and to provide an experimental database that can be used for the evaluation, modification, and validation of force field parameters that are needed to accurately represent protein adsorption behavior for molecular simulations.
PMCID: PMC3006061  PMID: 21073182
20.  Measuring bacterial cells size with AFM 
Brazilian Journal of Microbiology  2012;43(1):341-347.
Atomic Force Microscopy (AFM) can be used to obtain high-resolution topographical images of bacteria revealing surface details and cell integrity. During scanning however, the interactions between the AFM probe and the membrane results in distortion of the images. Such distortions or artifacts are the result of geometrical effects related to bacterial cell height, specimen curvature and the AFM probe geometry. The most common artifact in imaging is surface broadening, what can lead to errors in bacterial sizing. Several methods of correction have been proposed to compensate for these artifacts and in this study we describe a simple geometric model for the interaction between the tip (a pyramidal shaped AFM probe) and the bacterium (Escherichia coli JM-109 strain) to minimize the enlarging effect. Approaches to bacteria immobilization and examples of AFM images analysis are also described.
PMCID: PMC3768968  PMID: 24031837
Atomic force microscopy (AFM); Escherichia coli; cell dimensions; bacteria visualization
21.  Membrane Characterization by Microscopic and Scattering Methods: Multiscale Structure 
Membranes  2011;1(2):91-97.
Several microscopic and scattering techniques at different observation scales (from atomic to macroscopic) were used to characterize both surface and bulk properties of four new flat-sheet polyethersulfone (PES) membranes (10, 30, 100 and 300 kDa) and new 100 kDa hollow fibers (PVDF). Scanning Electron Microscopy (SEM) with “in lens” detection was used to obtain information on the pore sizes of the skin layers at the atomic scale. White Light Interferometry (WLI) and Atomic Force Microscopy (AFM) using different scales (for WLI: windows: 900 × 900 μm2 and 360 × 360 μm2; number of points: 1024; for AFM: windows: 50 × 50 μm2 and 5 × 5 μm2; number of points: 512) showed that the membrane roughness increases markedly with the observation scale and that there is a continuity between the different scan sizes for the determination of the RMS roughness. High angular resolution ellipsometric measurements were used to obtain the signature of each cut-off and the origin of the scattering was identified as coming from the membrane bulk.
PMCID: PMC4021929  PMID: 24957612
membrane; atomic force microscopy; white light interferometry; ellipsometry; roughness
22.  Relating the Physical Properties of Pseudomonas aeruginosa Lipopolysaccharides to Virulence by Atomic Force Microscopy▿ †  
Journal of Bacteriology  2010;193(5):1259-1266.
Lipopolysaccharides (LPS) are an important class of macromolecules that are components of the outer membrane of Gram-negative bacteria such as Pseudomonas aeruginosa. P. aeruginosa contains two different sugar chains, the homopolymer common antigen (A band) and the heteropolymer O antigen (B band), which impart serospecificity. The characteristics of LPS are generally assessed after isolation rather than in the context of whole bacteria. Here we used atomic force microscopy (AFM) to probe the physical properties of the LPS of P. aeruginosa strain PA103 (serogroup O11) in situ. This strain contains a mixture of long and very long polymers of O antigen, regulated by two different genes. For this analysis, we studied the wild-type strain and four mutants, ΔWzz1 (producing only very long LPS), ΔWzz2 (producing only long LPS), DΔM (with both the wzz1 and wzz2 genes deleted), and Wzy::GM (producing an LPS core oligosaccharide plus one unit of O antigen). Forces of adhesion between the LPS on these strains and the silicon nitride AFM tip were measured, and the Alexander and de Gennes model of steric repulsion between a flat surface and a polymer brush was used to calculate the LPS layer thickness (which we refer to as length), compressibility, and spacing between the individual molecules. LPS chains were longest for the wild-type strain and ΔWzz1, at 170.6 and 212.4 nm, respectively, and these values were not statistically significantly different from one another. Wzy::GM and DΔM have reduced LPS lengths, at 34.6 and 37.7 nm, respectively. Adhesion forces were not correlated with LPS length, but a relationship between adhesion force and bacterial pathogenicity was found in a mouse acute pneumonia model of infection. The adhesion forces with the AFM probe were lower for strains with LPS mutations, suggesting that the wild-type strain is optimized for maximal adhesion. Our research contributes to further understanding of the role of LPS in the adhesion and virulence of P. aeruginosa.
PMCID: PMC3067594  PMID: 21148734
23.  AFM-assisted fabrication of thiol SAM pattern with alternating quantified surface potential 
Nanoscale Research Letters  2011;6(1):185.
Thiol self-assembled monolayers (SAMs) are widely used in many nano- and bio-technology applications. We report a new approach to create and characterize a thiol SAMs micropattern with alternating charges on a flat gold-coated substrate using atomic force microscopy (AFM) and Kelvin probe force microscopy (KPFM). We produced SAMs-patterns made of alternating positively charged, negatively charged, and hydrophobic-terminated thiols by an automated AFM-assisted manipulation, or nanografting. We show that these thiol patterns possess only small topographical differences as revealed by AFM, and distinguished differences in surface potential (20-50 mV), revealed by KPFM. The pattern can be helpful in the development of biosensor technologies, specifically for selective binding of biomolecules based on charge and hydrophobicity, and serve as a model for creating surfaces with quantified alternating surface potential distribution.
PMCID: PMC3211238  PMID: 21711703
24.  Atomic force microscopy analysis of nanoparticles in non-ideal conditions 
Nanoscale Research Letters  2011;6(1):514.
Nanoparticles are often measured using atomic force microscopy or other scanning probe microscopy methods. For isolated nanoparticles on flat substrates, this is a relatively easy task. However, in real situations, we often need to analyze nanoparticles on rough substrates or nanoparticles that are not isolated. In this article, we present a simple model for realistic simulations of nanoparticle deposition and we employ this model for modeling nanoparticles on rough substrates. Different modeling conditions (coverage, relaxation after deposition) and convolution with different tip shapes are used to obtain a wide spectrum of virtual AFM nanoparticle images similar to those known from practice. Statistical parameters of nanoparticles are then analyzed using different data processing algorithms in order to show their systematic errors and to estimate uncertainties for atomic force microscopy analysis of nanoparticles under non-ideal conditions. It is shown that the elimination of user influence on the data processing algorithm is a key step for obtaining accurate results while analyzing nanoparticles measured in non-ideal conditions.
PMCID: PMC3212053  PMID: 21878120
25.  (2n × 1) Reconstructions of TiO2(011) Revealed by Noncontact Atomic Force Microscopy and Scanning Tunneling Microscopy 
We have used noncontact atomic force microscopy (NC-AFM) and scanning tunneling microscopy (STM) to study the rutile TiO2(011) surface. A series of (2n × 1) reconstructions were observed, including two types of (4 × 1) reconstruction. High-resolution NC-AFM and STM images indicate that the (4 × 1)-α phase has the same structural elements as the more widely reported (2 × 1) reconstruction. An array of analogous higher-order (2n × 1) reconstructions were also observed where n = 3–5. On the other hand, the (4 × 1)-β reconstruction seems to be a unique structure without higher-order analogues. A model is proposed for this structure that is also based on the (2 × 1) reconstruction but with additional microfacets of {111} character.
PMCID: PMC4191060  PMID: 25309642

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