Single molecule atomic force microscopy and force spectroscopy of chitosan

Atomic force microscopy (AFM) and AFM-based force spectroscopy was used to study the desorption of individual chitosan polymer chains from substrates with varying chemical composition. AFM images of chitosan adsorbed onto a flat mica substrate show elongated single strands or aggregated bundles. The aggregated state of the polymer is consistent with the high level of flexibility and mobility expected for a highly positively charged polymer strand. Conversely, the visualization of elongated strands indicated the presence of stabilizing interactions with the substrate. Surfaces with varying chemical composition (glass, self-assembled monolayer of mercaptoundecanoic acid/decanethiol and polytetrafluoroethylene (PTFE)) were probed with chitosan modified AFM tips and the corresponding desorption energies, calculated from plateau-like features, were attributed to the desorption of individual polymer strands. Desorption energies of 2.0±0.3×10(-20)J, 1.8±0.3×10(-20)J and 3.5±0.3×10(-20)J were obtained for glass, SAM of mercaptoundecanoic/dodecanethiol and PTFE, respectively. These single molecule level results can be used as a basis for investigating chitosan and chitosan-based materials for biomaterial applications.     

A combined vibrational sum frequency generation spectroscopy and atomic force microscopy study of sphingomyelin-cholesterol monolayers

A combination of vibrational sum frequency generation spectroscopy and atomic force microscopy is used to study the changes in morphology and conformational order in monolayers prepared from three natural sphingomyelin (SM) mixtures as a function of surface pressure and cholesterol concentration. The most homogeneous SM gave monolayers with well-ordered acyl chains and few gauche defects with relatively small effects of either increasing surface pressure or cholesterol addition. Heterogeneous SM mixtures with a mixture of acyl chain lengths or with significant fractions of unsaturated acyl chains had much larger contributions from gauche defects at low surface pressure and gave increasingly well-ordered monolayers as the surface pressure increased. They also showed substantial increases in lipid chain order after cholesterol addition. Overall, these results are consistent with the strong hydrogen bonding capacity of SM leading to well-ordered monolayers over a range of surface pressures. The changes in acyl chain order for natural SMs as a function of cholesterol are relevant to formation of sphingolipid-cholesterol enriched domains in cell membranes.     

Application of atomic force microscopy to the study of blown polyethylene films

Atomic force microscopy (AFM) has been used for the characterization of the surface topography and microstructure of polyethylene (PE) films with thickness of about 50 μm. Different compositions of the films were tested, including mixtures of LDPE fabricated with metallocene polyethylene (mPE). The characteristics of the fibrils and spherulites of the films have been observed by means of AFM without any preparation of the samples, allowing also differentiation of the amorphous and crystalline zones. A method is proposed for the quantification of the proportion of crystallinity based on the roughness of the films.     

Single molecule force spectroscopy on poly(vinyl alcohol) by atomic force microscopy

The elastic properties of poly(vinyl alcohol) (PVA) were investigated on the nanoscale using the new technique of single molecule force spectroscopy by atomic force microscopy (AFM). It was found that the elastic properties of PVA molecules scale linearly with their contour lengths. This finding corroborates that the deformation of individual PVA chains is measured. The force spectra of PVA show a kink at around 200 pN and cannot be fitted by an extended Langevin function. The deviation of the elastic behavior of PVA from a freely jointed chain model may indicate the presence of a suprastructure of PVA in NaCl solution.     

Local surface charge dissipation studied using force spectroscopy method of atomic force microscopy

We propose herein a method to study local surface charge dissipation in dielectric films using force spectroscopy technique of atomic force microscopy. By using a normalization procedure and considering an analytical expression of the tip‐sample interaction force, we could estimate the characteristic time decay of the dissipation process. This approach is completely independent of the atomic force microscopy tip geometry and considerably reduces the amount of experimental data needed for the calculation compared with other techniques. The feasibility of the method was demonstrated in a freshly cleaved mica surface, in which the local charge dissipation after cleavage followed approximately a first‐order exponential law with the characteristic time decay of approximately 7–8 min at 30% relative humidity (RH) and 2–3.5 min at 48% RH. Copyright © 2015 John Wiley & Sons, Ltd.     

Characterization of Langmuir-Blodgett organoclay films using X-ray reflectivity and atomic force microscopy

Monolayers of organoclay platelets were formed at the air/water interface using the Langmuir technique and were then investigated either by in situ or lifted onto Si wafers and studied ex situ, using X-ray reflectivity (XR) methods. The XR data showed that the surfactant molecules on the clay platelets formed a dense, self-assembled monolayer where the molecules were tilted at an angle of 35 degrees +/-6 degrees from the normal to the dry clay surface. The surfactant layers only covered a fraction of the clay platelet surface area, where the fractional surface coverage for the three clays studied (C6A, C15A, and C20A) was found to be 0.90, 0.86, and 0.73, respectively. These values were significantly higher than those estimated from the cation exchange capacity (CEC) values. Rather than being uniformly distributed, the surfactant was clustered in patchy regions, indicating that the surface of the clay platelets had both polar and non-polar segments. This heterogeneity confirmed the hypothesis which was previously invoked to explain the distribution of the clay platelets in melt mixed homopolymer and polymer blend nanocomposites.     

Acoustics and atomic force microscopy for the mechanical characterization of thin films

The science and technology of thin films require the development of nondestructive methods for their quantitative mechanical characterization with nanometric spatial resolution. High-frequency ultrasonic techniques—especially acoustic microscopy—and atomic force microscopy (AFM) have been demonstrated to represent versatile tools for developing such methods. In particular, in the last 15 years, the combination of AFM, which can probe the surface of a sample by applying ultralow loads (from micronewtons down to piconewtons) with a micromachined tip having an apex radius of a few nanometers, and ultrasonics techniques led researchers to develop some unique tools which allow one to perform not only spot measurements of the sample elastic modulus, but also to obtain both the qualitative imaging of mechanical properties and the quantitative mapping of the elastic modulus of the sample surface with nanometric lateral resolution. In the present review, firstly a brief overview of the main ultrasound-based techniques for thin film characterization is reported. Then, some of the ultrasonic AFM techniques are described, emphasizing their capability of retrieving maps of both the tip–sample contact stiffness and the sample elastic modulus. Although these techniques are less affected by the mechanical properties of the substrates than standard indentation tests, a method for the correction of the substrate effect in ultrathin films is reported in detail. Finally, by probing the mechanical properties of a small portion of the sample volume underneath the tip, we illustrate the techniques as tools for the qualitative and quantitative characterization of variations in the adhesion between a thin film and a buried interface, as well as for detecting subsurface defects, voids, cracks, and dislocations.     

Nanoscale degradation of polypyrrole films under oxidative stress: An atomic force microscopy study and review

Atomic force microscopy (AFM) is employed to monitor the surface morphology of polypyrrole (PPy) films grown on vitreous carbon substrates during the catalytic reduction of Cr(VI) to Cr(III). The morphology of freshly-prepared films depends on substrate characteristics. Upon reaction, uniform nodules of aggregated PPy clusters appear. No significant differences in surface morphology are found between its oxidized and reduced forms. Loss of catalytic activity after 8-9 oxidation/reduction cycles of exposure to the chromate solution (oxidation) and electrochemical recharging of the film at negative potentials (reduction) correlates well with the observed polymer film dissolution/detachment from the carbon substrate. Formation of well-defined circular features (PPy rings) at different stages leads to a model for the film degradation process that includes formation of Cl₂ gas inside the polymer matrix. In the final stages, the bulk of the film typically fractures and detaches from the electrode. A catalytically inactive, ultrathin PPy layer remains on the substrate even after prolonged exposure to the target solution. A review of techniques for the study of PPy aging/degradation is given.     

Correction of systematic errors in single-molecule force spectroscopy with polymeric tethers by atomic force microscopy

Single-molecule force spectroscopy has become a valuable tool for the investigation of intermolecular energy landscapes for a wide range of molecular associations. Atomic force microscopy (AFM) is often used as an experimental technique in these measurements, and the Bell-Evans model is commonly used in the statistical analysis of rupture forces. Most applications of the Bell-Evans model consider a constant loading rate of force applied to the intermolecular bond. The data analysis is often inconsistent because either the probe velocity or the apparent loading rate is being used as an independent parameter. These approaches provide different results when used in AFM-based experiments. Significant variations in results arise from the relative stiffness of the AFM force sensor in comparison with the stiffness of polymeric tethers that link the molecules under study to the solid surfaces. An analytical model presented here accounts for the systematic errors in force-spectroscopy parameters arising from the nonlinear loading induced by polymer tethers. The presented analytical model is based on the Bell-Evans model of the kinetics of forced dissociation and on the asymptotic models of tether stretching. The two most common data reduction procedures are analyzed, and analytical expressions for the systematic errors are provided. The model shows that the barrier width is underestimated and that the dissociation rate is significantly overestimated when force-spectroscopy data are analyzed without taking into account the elasticity of the polymeric tether. Systematic error estimates for asymptotic freely jointed chain and wormlike chain polymer models are given for comparison. The analytical model based on the asymptotic freely jointed chain stretching is employed to analyze and correct the results of the double-tether force-spectroscopy experiments of disjoining "hydrophobic bonds" between individual hexadecane molecules that are covalently tethered via poly(ethylene glycol) linkers of different lengths to the substrates and to the AFM probes. Application of the correction algorithm decreases the spread of the data from the mean value, which is particularly important for measurements of the dissociation rate, and increases the barrier width to 0.43 nm, which might be indicative of the theoretically predicted hydrophobic dewetting.     

Reverse engineering of an affinity-switchable molecular interaction characterized by atomic force microscopy single-molecule force spectroscopy

Tunable and switchable interaction between molecules is a key for regulation and control of cellular processes. The translation of the underlying physicochemical principles to synthetic and switchable functional entities and molecules that can mimic the corresponding molecular functions is called reverse molecular engineering. We quantitatively investigated autoinducer-regulated DNA-protein interaction in bacterial gene regulation processes with single atomic force microscopy (AFM) molecule force spectroscopy in vitro, and developed an artificial bistable molecular host-guest system that can be controlled and regulated by external signals (UV light exposure and thermal energy). The intermolecular binding functionality (affinity) and its reproducible and reversible switching has been proven by AFM force spectroscopy at the single-molecule level. This affinity-tunable optomechanical switch will allow novel applications with respect to molecular manipulation, nanoscale rewritable molecular memories, and/or artificial ion channels, which will serve for the controlled transport and release of ions and neutral compounds in the future.     

Combined atomic force microscopy and fluorescence correlation spectroscopy measurements to study the dynamical structure of interfacial fluids

We have studied the dynamic structure of thin (approximately a few nanometers) liquid films of a nearly spherical, nonpolar molecule tetrakis(2-ethylhexoxy)silane (TEHOS) by using a combination of atomic force microscopy (AFM) and fluorescence correlation spectroscopy (FCS). Ultra-sensitive interferometer-based AFM was used to determine the stiffness (force gradient) and the damping coefficient of the liquid film. The experiments show oscillations in the damping coefficient with a period of approximately 1 nm, which is consistent with the molecular dimension of TEHOS as well as previous X-ray reflectivity measurements. Additionally, we performed FCS experiments for direct determination of the molecular dynamics within the liquid film. From the fluctuation autocorrelation curve, we measured the translational diffusion of the probe molecule embedded within the fluid film formed on a solid substrate. The autocorrelation function was best fitted with two components, which indicate that the dynamics are heterogeneous in nature. However, the heterogeneity is not as pronounced as had been previously observed for molecularly thin liquid films sandwiched between two solid substrates.     

Probing the viscoelastic response of glassy polymer films using atomic force microscopy

The mechanical properties of glassy films and glass surfaces have been studied using an atomic force microscope (AFM) through various imaging modes and measuring methods. In this paper, we discuss the viscoelastic response of a glassy surface probed using an AFM. We analyzed the force-distance curves measured on a glassy film or a glassy surface at temperatures near the glass transition temperature, Tg, using a Burgers model. We found that the material's characteristics of reversible anelastic response and viscous creep can be extracted from a force-distance curve. Anelastic response shifts the repulsive force-distance curve while viscous creep strongly affects the slope of the repulsive force-distance curve. When coupled with capillary force, due to the condensation of a thin layer of liquid film at the tip-surface joint, the anelasticity and viscous creep can alter the curve significantly in the attractive region.     

Mechanical characterization of polymeric thin films by atomic force microscopy based techniques

Polymeric thin films have been awakening continuous and growing interest for application in nanotechnology. For such applications, the assessment of their (nano)mechanical properties is a key issue, since they may dramatically vary between the bulk and the thin film state, even for the same polymer. Therefore, techniques are required for the in situ characterization of mechanical properties of thin films that must be nondestructive or only minimally destructive. Also, they must also be able to probe nanometer-thick ultrathin films and layers and capable of imaging the mechanical properties of the sample with nanometer lateral resolution, since, for instance, at these scales blends or copolymers are not uniform, their phases being separated. Atomic force microscopy (AFM) has been proposed as a tool for the development of a number of techniques that match such requirements. In this review, we describe the state of the art of the main AFM-based methods for qualitative and quantitative single-point measurements and imaging of mechanical properties of polymeric thin films, illustrating their specific merits and limitations.     

High-resolution atomic force microscopy study of hexaglycylamide epitaxial structures on graphite

Two types of hexaglycylamide (HGA) epitaxial lamellar structures coexisting on the surface of highly oriented pyrolytic graphite (HOPG) exposed to water solutions were studied by high-resolution atomic force microscopy (AFM). Lamellae are distinguished by growth direction and by morphology. The lamellae of the first type (L1) produced by depositions from more dilute solutions are close-packed with a period of ～5.2 nm, twice the HGA molecular length, and form highly ordered domains morphologically similar to the lamellar domains of alkanes. The less-ordered lamellae of the second type (L2) appear at intermediate and large HGA concentrations and demonstrate variable lamellar width, morphological diversity, and a tendency to merge. The interlamellar separation in the domains of close-packed L2 lamellae varies with the discrete increment ～2.5 nm; the most frequently observed value is ～7.5-8.0 nm corresponding to the triple HGA molecular length. The growth directions of lamellae of each type have sixfold rotational symmetry indicating epitaxy with graphite; however, the rosettes of L1 and L2 lamellae orientations are misaligned by 30°. The molecular modeling of possible HGA epitaxial packing arrangements on graphite and their classification have been conducted, and the energetically preferable structures are selected. On this basis, the structural models of the L1 and L2 lamellae are proposed explaining the experimentally observed peculiarities as follows: (1) the L1 and L2 lamellae are respectively parallel and antiparallel β-sheets with two HGA molecules in the unit cell oriented normally to the lamellae boundaries, (2) HGA molecules in L1 and L2 lamellae have different orientations with respect to the graphite lattice, respectively along the directions <1120> and <1010>, (3) L1 lamella is the assembly of two hydrogen-bonded parallel β-sheets oriented head-to-head, (4) L2 lamellae are assemblies of several molecular rows (antiparallel β-sheets) cross-linked by hydrogen bonds. The AFM observations indicate that the covering of the hydrophobic graphite by the dense, closely packed, well-ordered monolayers of hydrophilic oligopeptide is possible.     

Atomic force spectroscopy on poly(o-ethoxyaniline) nanostructured films: sensing nonspecific interactions.

Atomic force spectroscopy (AFS) was used to measure interaction forces between the tip and nanostructured layers of poly(o-ethoxyaniline) (POEA) in pure water and CuSO4 solutions. When the tip approach and retraction were carried out at low speeds, POEA chains could be physisorbed onto the Si3N4 tip via nonspecific interactions. We conjecture that while detaching, POEA chains were stretched and the estimated chain lengths were consistent with the expected values from the measured POEA molecular weight. The effects from POEA doping could be investigated directly by performing AFS measurements in a liquid cell, with the POEA film exposed to liquids of distinct pH values. For pH > or = 6.0, the force curves normally displayed an attractive region for POEA, but at lower pH values-where POEA is protonated-the repulsive double-layer forces dominated. Measurements in the liquid cell could be further exploited to investigate how the film morphology and the force curve are affected when impurities are deliberately introduced in the liquid. The shape of the force curves and the film morphology depended on the concentration of heavy metal in the liquid cell. AFS may therefore be used to study the interaction between film and analyte, with important implications for the understanding of mechanisms governing the sensing ability of taste sensors.     

Comparative characterisation by atomic force microscopy and ellipsometry of soft and solid thin films

Ellipsometry and atomic force microscopy (AFM) were used to study the film thickness and the surface roughness of both ‘soft’ and solid thin films. ‘Soft’ polymer thin films of polystyrene and poly(styrene–ethylene/butylene–styrene) block copolymer were prepared by spin‐coating onto planar silicon wafers. Ellipsometric parameters were fitted by the Cauchy approach using a two‐layer model with planar boundaries between the layers. The smooth surfaces of the prepared polymer films were confirmed by AFM. There is good agreement between AFM and ellipsometry in the 80–130 nm thickness range. Semiconductor surfaces (Si) obtained by anisotropic chemical etching were investigated as an example of a randomly rough surface. To define roughness parameters by ellipsometry, the top rough layers were treated as thin films according to the Bruggeman effective medium approximation (BEMA). Surface roughness values measured by AFM and ellipsometry show the same tendency of increasing roughness with increased etching time, although AFM results depend on the used window size. The combined use of both methods appears to offer the most comprehensive route to quantitative surface roughness characterisation of solid films. Copyright © 2007 John Wiley & Sons, Ltd.     

Investigation of Ti/CuO interface by X-ray photoelectron spectroscopy and atomic force microscopy

The Ti/CuO interface has been studied by the techniques of X-ray photoelectron spectroscopy and atomic force microscopy. Thin films of titanium were deposited on a CuO substrate at room temperature by the e-beam technique. The photoelectron spectra of titanium and copper were found to exhibit significant chemical interaction at the interface. The titanium overlayer was observed to get oxidized to TiO₂, while the CuO was observed to get reduced to elemental copper. This chemical interaction was observed to occur until a thickness of 7 nm of the titanium overlayer. For thicknesses greater than this value, the presence of unreacted titanium in the sample was detected. Barrier characteristics at the Ti/CuO interface were also carried out for substrate temperatures of 300°C, 400°C, 500°C, and 600°C as a function of the titanium overlayer thickness. A linear trend in the barrier thickness of the overlayer was observed between 400°C and 600°C substrate temperatures. The atomic force microscopy micrographs of the unannealed samples depicted layer-by-layer growth of elemental titanium on copper. At the Ti/CuO interface in such samples, the micrographs exhibited island formation of TiO₂ corresponding to the Volmer-Weber growth model. This formation has been interpreted as the relaxation in the strain energy. The percentage coverage of the underlying substrate by the TiO₂ islands showed a linear trend for the thicknesses of the titanium overlayer investigated. The average size of these islands also showed a linear trend as a function of the thickness of the overlayer.     

Atomic force microscopy study of the interaction of DNA and nanostructured beta-Gallia rutile

The ability to attach DNA molecules to solid planar substrates is desired for imaging the molecule and for building DNA-mediated nanostructures. The deposition of DNA on [001] rutile and beta-gallia rutile (BGR) substrates from buffer solutions containing various divalent cations was studied using tapping mode atomic force microscopy (AFM). beta-Gallia rutile intergrowths were prepared by spin-coating gallium isopropoxide onto [001]-oriented TiO2 single-crystal slabs and heating above 1350 degrees C for >24 h, resulting in the formation of intergrowth lines along the {210} planes in the parent rutile structure. Rutile and BGR intergrowth substrates were exposed to various buffered solutions containing DNA and the following divalent cations: Ca(II), Co(II), Cu(II), Fe(II), Mg(II), Mn(II), Ni(II), and Zn(II). Among all the cations examined, only Ni(II) resulted in the attachment of DNA on the rutile surfaces. DNA attachment to BGR surfaces was strong enough to allow AFM imaging when the deposition buffer contained one of the following cations: Co(II), Mg(II), Mn(II), Ni(II), and Zn(II). For all of these cations, DNA attachment occurred preferentially, but not exclusively, along BGR intergrowth lines. When buffers without cation additions and those containing Ca(II), Cu(II), and Fe(II) were used, DNA failed to bind the BGR surfaces strongly enough to allow AFM imaging. The mechanism(s) by which DNA attaches to the BGR surface is (are) not well understood but may involve the incorporation of divalent cations at the tunnel sites of the BGR intergrowths.     

In situ atomic force microscopy of zeolite A dissolution

In the present study, the {100} surface of zeolite A was exposed to a range of solutions and the response was monitored in real-time by means of atomic force microscopy (AFM). The zeolite dissolves by a well-defined layer process that is characterised by uncorrelated dissolution of units that are structurally unconnected and terrace retreat when building units are inter-connected. This process was observed to be coupled with the formation of nano-squares that are stabilized at the zeolite surface for a period before complete dissolution. Theoretical work suggests that three terminating structures are central to understanding the dissolution mechanism. Stripping the surface of the secondary building unit, the single 4-ring, is predicted to be a rate-determining step in dissolution, but this process occurs by removing monomeric rather than oligomeric units.