Abstract Atomic Force Microscopy Images Research Articles

AI-based atomic force microscopy image analysis allows to predict electrochemical impedance spectra of defects in tethered bilayer membranes

Atomic force microscopy (AFM) image analysis of supported bilayers, such as tethered bilayer membranes (tBLMs) can reveal the nature of the membrane damage by pore-forming proteins and predict the electrochemical impedance spectroscopy (EIS) response of such objects. However, automated analysis involving pore detection in such images is often non-trivial and can require AI-based object detection techniques. The specific object-detection algorithm we used to determine the defect coordinates in real AFM images was a convolutional neural network (CNN). Defect coordinates allow to predict the EIS response of tBLMs populated by the pore-forming toxins using finite element analysis (FEA) modeling. We tested if the accuracy of the CNN algorithm affected the EIS spectral features sensitive to defect densities and other physical parameters of tBLMs. We found that the EIS spectra can be predicted sufficiently well, however, systematic errors of characteristic spectral points were observed and need to be taken into account. Importantly, the comparison of predicted EIS curves with experimental ones allowed to estimate important physical parameters of tBLMs such as the specific resistance of submembrane reservoir. This reservoir separates phospholipid bilayer from the solid support. We found that the specific resistance of the reservoir amounts to 10^{4.25 pm 0.10}Omega cdot cm which is approximately two orders of a magnitude higher compared to the specific resistance of the buffer bathing tBLMs studied in this work. We hypothesize that such effect may be related in part due to decreased concentration of ionic carriers in the submembrane due to decreased relative dielectric permittivity in this region.

Characterizing AFM Tip Lateral Positioning Variability Through Non-Vector Space Control-Based Nanometrology

Atomic force microscopy (AFM) based nanotechnology has been widely implemented in various fields for decades in light of its overwhelming advantages, such as nanometer spatial resolution, adaptability to liquid ambient, and various nanomechanical/electrical metrological approaches. It is noted that though AFM possesses imaging capability up to nanometer resolution, it is hard to achieve nanometer level positioning precision due to the existing system variability, especially the thermal drift, which distorts AFM images through relatively long capturing time. Since an AFM image is typically utilized as a global reference map to navigate its tip to the desired locations for precise measurement and manipulation, the system variability distorted image will definitely diversify the experimental results. Therefore, it is necessary to characterize the positioning variability for better experimental results evaluation and decision-making. Although various approaches were proposed to evaluate AFM positioning error, to our best knowledge, there is little research about characterizing its positioning variability precisely and systematically. In this study, we present a universal metrological approach to quantitatively measure AFM tip locating variability by developing a featureless spiral local scan strategy together with the non-vector space (NVS) navigation approach. As a demonstration, the proposed nanometrology was conducted on a specific AFM platform to unravel its positioning property.

Experimental analysis of rough surface topography and modifying the humidity effect in AFM images to improve the topography quality

Atomic force microscopy (AFM) is one of the most powerful tools for imaging surfaces at the nano-scale. The existence of disruptive forces in the output images that are obtained from AFM in a moist environment causes low-quality images at the nano-scale. Therefore, to enhance the quality of topography, eliminating the effect of these forces is of great importance. For this purpose, increasing accuracy in modeling and simulation of vibration behavior of AFM piezoelectric micro-cantilever (MC) and its comparison with experimental tests is important to improve the quality of images. Using modified couple stress (MCS) and classical theory, this article addresses inhomogeneous four-layered AFM MC considering Timoshenko and Euler-Bernoulli beam models in 2D modeling. In 3D modeling by using the COMSOL software, cantilever frequency and time responses were obtained and compared with different theories including classical and MCS. Laboratory tests were conducted not only to validate results but also to evaluate the effect of humidity on topography image quality in the amplitude mode. Furthermore, using simulation results based on the Timoshenko beam theory and MC behavior prediction, the capillary effect which causes noise in experimental images was filtered and the image quality which is taken by the AFM is improved.

Atomic force and scanning near-field optical microscopy study of carbocyanine dye J-aggregates

Atomic force microscopy (AFM) and scanning near-field optical microscopy (SNOM) imaging of fibrillar J-aggregates of carbocyanine dye 3,3′-bis(γ-sulfopropyl)-5,5′,6,6′-tetrachloro-1,1′-dioctylbenzimidacarbocyanine (C8S3) is performed on mica and glass substrates at room temperature. High resolution AFM imaging reveal that thick fibrils consist of narrow sub-fibrils. A model of an elementary fibril with a cross section 3 × 4 nm2 in size is proposed. The SNOM measurements made in the contact mode indicate the high mechanical strength of the fibrillar J-aggregates.

BIAXIALLY ORIENTED POLYPROPYLENE FILM FOR AFM TIP CHARACTERIZATION

Atomic force microscopy (AFM) investigating the sample morphology is the process of direct interaction between tip and surface features. The geometry of probe tip is a determining factor in correcting AFM images distorted by tip size itself. A quantitative knowledge of the current tip shape is needed to improve the reliability of AFM images. The biaxially oriented polypropylene (BOPP) film was fabricated and used as tip characterizer to estimate the morphology of AFM Si tip based on the blind reconstruction algorithm. The surface of the BOPP film was covered by nanometer-scale-sized structures which ensure that the tip profile can be determined accurately. Without independent knowledge of the sample, the three-dimensional (3D) shape of Si probe tip was obtained with high aspect ratio. BOPP film is not only a simple, cheap material but also a soft one which can also avoid tip damage in scanning. It was demonstrated reliable and suitable for tip characterization.

Reconstruction of Atomic Force Microscope Image Using Estimated Tip Shape from Impulse Response Technique

Atomic force microscopy (AFM) is the useful tool for measuring the micro fabricated devices. Measuring sample surface with AFM, tip-induced distortions are serious problem for accurate measurement. In order to overcome this problem, many studies are reported to reconstruct surface image to original sample surface. We studied method of reconstruction technique of the AFM image by using estimated 3-D tip shape from the impulse response technique. To reconstruct the image of the sample surface, we used a morphological reconstruction process. We demonstrated how to obtain the 3-D tip from the using impulse response technique and then how to reconstruct the 3-D AFM image using estimated tip shape.

Investigation of the structural organization of cationic nanoemulsion/antisense oligonucleotide complexes

Atomic force microscopy image analysis and energy dispersive X-ray diffraction experiments were used to investigate the structural organization of cationic nanoemulsion/oligonucleotide complexes. Oligonucleotides targeting topoisomerase II gene were adsorbed on cationic nanoemulsions obtained by means of spontaneous emulsification procedure. Topographical analysis by atomic force microscopy allowed the observation of the nanoemulsion/oligonucleotide complexes through three-dimensional high-resolution images. Flattening of the oil droplets was observed, which was reduced in the complexes obtained at high amount of adsorbed oligonucleotides. In such conditions, complexes exhibit droplet size in the 600nm range. The oligonucleotides molecules were detected on the surface of the droplets, preventing their fusion during aggregation. A lamellar structure organization was identified by energy dispersive X-ray diffraction experiments. The presence of the nucleic acid molecules led to a disorganization of the lipid arrangement and an expansion in the lattice spacing, which was proportional to the amount of oligonucleotides added.

Imaging and Measuring the Molecular Force of Lymphoma Pathological Cells Using Atomic Force Microscopy

Atomic force microscopy (AFM) provides a new technology to visualize the cellular topography and quantify the molecular interactions at nanometer spatial resolution. In this work, AFM was used to image the cellular topography and measure the molecular force of pathological cells from B-cell lymphoma patients. After the fluorescence staining, cancer cells were recognized by their special morphological features and then the detailed topography was visualized by AFM imaging. The AFM images showed that cancer cells were much rougher than healthy cells. CD20 is a surface marker of B cells and rituximab is a monoclonal antibody against CD20. To measure the CD20-rituximab interaction forces, the polyethylene glycol (PEG) linker was used to link rituximab onto the AFM tip and the verification experiments of the functionalized probe indicated that rituximab molecules were successfully linked onto the AFM tip. The CD20-rituximab interaction forces were measured on about 20 pathological cells and the force measurement results indicated the CD20-rituximab binding forces were mainly in the range of 110-120 pN and 130-140 pN. These results can improve our understanding of the topography and molecular force of lymphoma pathological cells.

Localization and force analysis at the single virus particle level using atomic force microscopy

Atomic force microscopy (AFM) is a vital instrument in nanobiotechnology. In this study, we developed a method that enables AFM to simultaneously measure specific unbinding force and map the viral glycoprotein at the single virus particle level. The average diameter of virus particles from AFM images and the specificity between the viral surface antigen and antibody probe were integrated to design a three-stage method that sets the measuring area to a single virus particle before obtaining the force measurements, where the influenza virus was used as the object of measurements. Based on the purposed method and performed analysis, several findings can be derived from the results. The mean unbinding force of a single virus particle can be quantified, and no significant difference exists in this value among virus particles. Furthermore, the repeatability of the proposed method is demonstrated. The force mapping images reveal that the distributions of surface viral antigens recognized by antibody probe were dispersed on the whole surface of individual virus particles under the proposed method and experimental criteria; meanwhile, the binding probabilities are similar among particles. This approach can be easily applied to most AFM systems without specific components or configurations. These results help understand the force-based analysis at the single virus particle level, and therefore, can reinforce the capability of AFM to investigate a specific type of viral surface protein and its distributions.

Characterization of Surface Coats of Bacterial Spores with Atomic Force Microscopy and Wavelets

Atomic force microscopy images of the surface coatings of bacterial spores can be analyzed by wavelet analysis to rapidly determine the characteristic morphology of the spore coat. The identification of bacterial spores in the environment is an important problem for preventing disease, identifying bacterial contamination of air, water, and soil, and evaluating the effects of chemicals on bacteria. In this work, we analyze AFM data of the native surface topography and ultra structure of spore coats of native Bacillus atrophaeus and Bacillus thuringiensis. We show that if the analysis of the images is done as a function of the rotation angle of the image observation, the morphology and the characteristic sizes of the bacterial coats can be accurately analyzed. The technique has potential to rapidly identify bacterial spores and viruses if a library of images of various bacteria are obtained.

Selective image analysis for roughness characterisation

Atomic force microscopy images supply precise information on real surface topography, but an automated method allowing accurate description of roughness through an asperity based approach is necessary for tribological applications. The present paper describes an original selective image analysis algorithm enabling confident indexation and analysis of surface asperities. First, the original image undergoes noise filtering followed by edge detection and thresholding. In the resulting binary image, clusters of asperities are separated by a selective treatment depending on objects' size and convexity. Finally, a regrowth of identified objects is applied to ensure their accurate three-dimensional shape estimation using a second order polynomial least square approximation. The identification performance of the method is hereby verified on an artificial image and illustrated with surface images of sputtered gold thin film alone and with an additional layer of ruthenium.

Surface and optical property modifications of self-assembled CdTe/ZnTe quantum dots caused by thermal treatment

Atomic force microscopy images showed that the size of the CdTe quantum dots (QDs) slightly increased with increasing annealing temperature up to 350 °C. Photoluminescence spectra showed that the excitonic peak corresponding to the interband transition from the ground electronic subband to the ground heavy-hole band (E1–HH1) in the CdTe/ZnTe QDs annealed at 350 °C was shifted to lower energy compared with that in as-grown CdTe/ZnTe QDs. The full width at half-maximum of the E1–HH1 transition peak in the CdTe/ZnTe QDs annealed at 350 °C decreased resulting from the improvement of the crystallinity for the annealed CdTe QDs.

AFM characterization of rabbit spermatozoa

Atomic force microscopy (AFM) has been applied for determining the topological and structural features of rabbit spermatozoa. Fresh ejaculated spermatozoa were adsorbed passively onto a silicon slide or by motility from suspension onto a poly(L-lysine)-coated glass coverslip and then imaged in air and in buffer saline, respectively. AFM images clearly highlighted many details of spermatozoa head, neck, and tail. Distinct features were observed in the plasmatic membrane of spermatozoa. In particular, head topography easily recognized the acrosome, equatorial segment, equatorial subsegment, and postacrosome regions. Moreover, AFM images revealed the presence of double belt of invaginations around the spermatozoa head, at the boundary between equatorial subsegment and postacrosome regions. All together, the collected AFM images clearly defined a detailed map of spermatozoa morphology while giving some hints on the internal structure.

An easy method to discover cell membrane antigen with atomic force microscopy

Atomic force microscopy (AFM) obtains a high resolution at nanometer. In the current study, we found that, before and after adding CD34, CD44 or CD29 antibodies, the AFM images of mouse adipose tissue derived mesenchymal stem cells (ADMSCs) had significant changes in both cell morphous (shape) and the average roughness of cell membrane. However, there was no significant difference in the cell shape and the average roughness of cell membrane, after adding CD45 or CD144 antibodies. Therefore, we could use the AFM to scan cell shape or to calculate the average roughness changes of cell membrane to analyze the existence of cell membrane antigen qualitatively.

AFM Analysis of C60 and a Poly(amido amine) Dendrimer-C60 Nanoconjugate

Atomic force microscopy (AFM) was used to study the nanoscopic structure and topography of buckminsterfullerene (C60) and a conjugate of C60 with generation four, amine-terminated, poly(amido amine) dendrimer (PAMAM-G4). The conjugate contains a PAMAM-G4 core and C60 shell formed by reacting PAMAM-G4 with an excess of C60. Fractal patterns of C60 were observed in nanoscopic AFM images when solutions of different concentrations of C60 in pyridine or toluene were dried at room temperature. In contrast, no fractal patterns were detected in the AFM images of the dendrimer-C60 nanoconjugate, prepared from pyridine solution in a similar manner. Thus, the C60-shell alone is not sufficient to impart the same fractal patterns on the conjugate.

Investigation of Sisal Fibers by Atomic Force Microscopy: Morphological and Adhesive Characteristics

Atomic force microscopy (AFM) was used to study the nanoscale surface chemistry and morphological changes caused by chemical treatment of sisal fibers. Scanning Electron Microscopy (SEM) micrographs indicated that sisal in natura (bundle of fibers) is formed by fibers with diameters of approximately 10 microm. AFM images showed that these fibers consist of microfibrils with diameters varying from 250 to 600 nm, which are made up of nanofibrils of ca. 20 nm in diameter. The adhesion force (pull-off force) between the AFM tip and the fibers surface increased after benzylation, pointing to a decrease in the polar groups on the sisal fiber. The adhesion map measured over a scan range of 3 microm was heterogeneous in samples treated with 40% NaOH and the low adhesion sites disappeared after benzylation. Using an established mathematical model, it was possible to evaluate the increase in adhesion work and consequently in the interaction between the AFM tip and sisal fibers. These results indicated that AFM can detect heterogeneity in the wettability of sisal fibers with nanometer resolution and can be applied in the study of fiber-matrix adhesion in polymer composites.

Atomic Force Microscopy in Analysis of Rubber Materials

Abstract Atomic force microscopy (AFM) and electric force microscopy (EFM) have been applied for compositional mapping of a number of elastomers and related multicomponent materials. Several aspects of optimizing AFM experiments on polymers are discussed. AFM images revealed changes of EPDM morphology caused by crosslinking and by loading with fillers [carbon black (CB) and silica particles] and oil. It was shown that the morphology of isotactic polypropylene (iPP)/EPDM vulcanizates, which were studied with AFM and EFM, depends on the ratio of components, degree of cure and processing conditions. Diffusion of oil from the elastomer component to the matrix is evidenced in the AFM images. Selective distribution of CB in the iPP matrix is responsible for the electric conductivity of the thermoplastic vulcanizate.

Study of the surfaces of CVD-WO3 films, by atomic force microscopy and spectroscopic ellipsometry

Atomic force microscopy imaging of chemical vapor deposition WO3 films reveals the presence of domed crystallites that resemble the florets of cauliflowers with a rough surface texture. Annealing at 400 °C and above leads to further surface roughening, with estimated root mean square roughness values of 40–50 nm. Spectroscopic ellipsometry analysis shows that the surface layer becomes thicker with increasing oxygen flow rate during film deposition. This layer is predominantly amorphous for as-deposited films, and predominantly crystalline after annealing.

Atomic force microscopy of scratch damage in polypropylene

Atomic force microscopy (AFM) in tapping mode has been used to characterise surface damage on deformed polypropylenes induced during a scratch test. Atomic force micrographs revealed differences in microstructures that could be used to predict the deformation resistance of two different types of polypropylene. The undeformed surface of the two types of polypropylene (identified as polypropylene-L and polypropylene-R) was characterised by differences in arrangement (regular or irregular) of fibrils depending on their melt flow conditions. Polypropylene-L is a polymer with longer chains and with restricted flow, whereas polypropylene-R has shorter chains obtained by controlled rheology. The microfibrils in undeformed polypropylene-L bend, forming raised surface features of height in the region of 10- 50 nm. In comparison to polypropylene-L, the microfibrils in undeformed polypropylene-R exhibited surface features of relatively lower height (10 - 20 nm). 30 × 30 nm scan AFM images provided details of microfibrils containing chains of molecules of ~0.5 nm wide. Surface deformation induced by the scratch resulted in the formation of scratch tracks characterised by regions of quasi-periodic (consecutive) cracking. This type of deformation is attributed to higher applied loads or to higher contact strains. This is particularly important in semicrystalline polymers, where there is partial reorganisation of microstructure on the application of surface stresses because of their viscoelastic properties. Atomic force micrographs of mechanically deformed polypropylene-L and polypropylene-R at a scan size of 1 × 1 μm indicated a lesser amount of reorganisation of microstructure in polypropylene-L as compared with polypropylene-R. Surface profiles and section analysis of the AFM micrographs suggested that polypropylene-R is more scratch resistant in comparison to polypropylene-L under identical scratch test conditions, consistent with Raman spectroscopy observations of tensile deformed polypropylene.

Elastic Recovery Measurements Performed by Atomic Force Microscopy and Standard Nanoindentation on a Co(10.1) Monocrystal

Atomic force microscopy (AFM) nanoindentation experiments were performed on a Co(10.1) monocrystal. Using AFM line scans, we deduced the elastic recovery, which is an intrinsic parameter of the studied material. The comparison of these elastic recovery values with those calculated by standard nanoindentation shows a fair agreement for forces higher than 400 μN with an important discrepancy for lower forces. This difference is attributed to tip shape effects and to the AFM cantilever elastic deformation. Furthermore, the material hardness was measured from AFM images of the imprint by considering the lateral dimension L. In this case, the obtained values are practically independent from the applied load. Moreover, a simple model based on geometrical considerations is proposed to correct hardness values calculated from the residual depth.