Advances In Atomic Force Microscopy Research Articles

Advances in Atomic Force Microscopy: Weakly Perturbative Imaging of the Interfacial Water.

The structure and dynamics of interfacial water, determined by the water-interface interactions, are important for a wide range of applied fields and natural processes, such as water diffusion (Kim et al., 2013), electrochemistry (Markovic, 2013), heterogeneous catalysis (Over et al., 2000), and lubrication (Zilibotti et al., 2013). The precise understanding of water-interface interactions largely relies on the development of atomic-scale experimental techniques (Guo et al., 2014) and computational methods (Hapala et al., 2014b). Scanning probe microscopy has been extensively applied to probe interfacial water in many interdisciplinary fields (Ichii et al., 2012; Shiotari and Sugimoto, 2017; Peng et al., 2018a). In this perspective, we review the recent progress in the noncontact atomic force microscopy (nc-AFM) imaging and AFM simulation techniques and discuss how the newly developed techniques are applied to study the properties of interfacial water. The nc-AFM with the quadrupole-like CO-terminated tip can achieve ultrahigh-resolution imaging of the interfacial water on different surfaces, trace the reconstruction of H-bonding network and determine the intrinsic structures of the weakly bonded water clusters and even their metastable states. In the end, we present an outlook on the directions of future AFM studies of interfacial water as well as the challenges faced by this field.

Use of Atomic Force Microscopy to Measure Mechanical Properties and Turgor Pressure of Plant Cells and Plant Tissues.

We present here the use of atomic force microscopy to indent plant tissues and recover its mechanical properties. Using two different microscopes in indentation mode, we show how to measure an elastic modulus and use it to evaluate cell wall mechanical properties. In addition, we also explain how to evaluate turgor pressure. The main advantages of atomic force microscopy are that it is non-invasive, relatively rapid (5~20 min), and that virtually any type of living plant tissue that is superficially flat can be analyzed without the need for treatment. The resolution can be very good, depending on the tip size and on the number of measurements per unit area. One limitation of this method is that it only gives direct access to the superficial cell layer.

Tunable Hybrid Biopolymeric Hydrogel Scaffolds Based on Atomic Force Microscopy Characterizations for Tissue Engineering.

Developing adequate biomaterials to engineer cell-scaffold interactions has become a promising way for physically regulating the biological behaviors of cells in the field of tissue engineering. Biopolymeric hydrogels have shown great merits as cellular scaffolds due to their biocompatible and biodegradable characteristics. In particular, the advent of atomic force microscopy (AFM) provides a powerful tool for characterizing native specimens at the micro/nanoscale, but utilizing AFM to investigate the detailed structures and properties of hydrogel scaffolds has been still scarce. In this paper, hybrid natural biopolymers are used to form hydrogel scaffolds which exhibit tunable structural and mechanical properties characterized by AFM peak force tapping imaging, and the applications of the formed hydrogel scaffolds in tissue engineering are studied. AFM morphological images showed that the cross-linking reactions of sodium alginate and gum arabic via calcium cations yielded the porous hydrogel scaffolds. By altering the component ratios, AFM mechanical images showed that the porous and mechanical properties (Young's modulus and adhesion force) of the hydrogel scaffolds were tunable. Next, the nanoscale structural and mechanical dynamics of the fabricated hydrogel scaffolds during the degradation process were revealed by AFM peak force tapping imaging. The experimental results on three different types of cells showed that the fabricated hydrogel scaffolds facilitate the formation of cellular spheroids. The research provides a novel idea to design tunable hydrogel scaffolds based on AFM characterizations for investigating cell-scaffold interactions, which will have potential impacts on tissue engineering.

Atomistic Simulation of Atomic Force Microscopy Imaging of Hydration Layers on Calcite, Dolomite, and Magnesite Surfaces

Advances in atomic force microscopy (AFM) in water have enabled the study of hydration layer structures on crystal surfaces, and in a recent study on dolomite (CaMg(CO3)2), chemical sensitivity was...

Measurement and analysis of cellular viscoelastic properties using atomic force microscopy

Cell mechanics plays an important role in cellular physiological and pathological processes. During the formation and progression of tumors, the alterations in the mechanics of cancerous cells and tumor micro-environment promote the growth and migration of cancerous cells. Hence, investigating cell mechanics is of crucial significance in understanding the underlying mechanisms regulating life activities and diseases. The advent of atomic force microscopy (AFM) provides a powerful tool for detecting the mechanical properties of single cells. Compared with other single-cell mechanical analysis techniques, the advantage of AFM is that AFM is able to simultaneously obtain the topography and mechanics of cells, which is particularly useful for investigating the correlation between cell structures and cell mechanics. The biomedical applications of AFM in single-cell mechanics provide considerable novel insights into how cell and tissue mechanics affect tumor development and metastasis, contributing much to the communities of biomechanics and biophysics. However, current AFM single-cell mechanical experiments are commonly performed on measuring the elastic properties of cells and studies about the viscoelastic properties of cells are still scarce. In this work, AFM was utilized to measure and analyze the viscoelastic properties of cells. First, the detailed procedure of detecting the viscoelasticity of cells based on AFM indentation technique was established. AFM probe was controlled to perform approach-dwell-retract movement on cells in the vertical direction. During the approach-dwell-retract process, the deflection of AFM cantilever versus time was recorded, which yielded the force-time (F-T) curves. The original F-T curves were then normalized and fitted by two-order Maxwell model, which gave two cellular relaxation times (the first relaxation time τ 1 and the second relaxation time τ 2). The fitting results showed that the theoretical curve matched the experimental relaxation curve well, indicating that the two-order Maxwell model was suited for characterizing the relaxation behaviors of cells. Second, the established procedure was used to measure the relaxation time of six different types of cells, including mammalian adherent cells, mammalian suspended cells, normal cells, cancerous cells, cell lines cultured in vitro and primary cells prepared from bone marrow and peripheral blood of healthy volunteers. The fitting curves were consistent with the experimental relaxation curves for all of the six types of cells used here, indicating the effectiveness of the presented method for detecting the viscoelastic properties of cells. Besides, the results showed the various cellular relaxation times for the different types of cells. The statistical results showed that the first cellular relaxation times were in the range 0.01−0.03 s, which might correspond to the behaviors of cytoplasm. The second cellular relaxation time was in the range 0.2−0.4 s, which might correspond to the behaviors of the cytoskeleton. Finally, regression analysis was performed on the measured cellular relaxation times, showing the linear relationship between the first cellular relaxation time τ 1 and the second cellular relaxation time τ 2, and the regression coefficients were variable for different types of cells. The research improves our understanding of cellular viscoelasticity and also provides a novel idea to measure the viscoelastic properties of cells, which will have potential impacts on cell mechanics and biomedicine.

Composite Nanostructures and Adhesion Analysis of Natural Plant Hydrogels Investigated by Atomic Force Microscopy.

Hydrogels designed by biomimetism and bioinspiration have been considered as a promising biomaterial for biomedical applications due to their biocompatible characteristics. Though biomimetic or bioinspired hydrogels have been widely studied by biochemical analysis, so far the detailed organizations and properties of native natural hydrogels are still not fully understood. The advent of atomic force microscopy (AFM) provides a potent tool for probing the biological samples in their living states with nanometer spatial resolution, which offers new possibilities for addressing the biological and material issues at the nanoscale. In this paper, AFM was utilized to characterize the structures and adhesive properties of the natural hydrogels produced by the carnivorous plant sundew. AFM morphological images of the mucilage secreted by three types of sundew show the composite nanostructures of the mucilage (e.g., nanoparticles, nanofibers, and porous polymeric networks), which are correlated with the mucilage's adhesive features revealed by AFM force measurements and peak force tapping mechanical imaging. The research demonstrates the prominent capabilities of AFM in characterizing the nanoscopic structures and properties of natural hydrogels with unprecedented spatial resolution, which is useful for understanding the underlying mechanisms guiding the behaviors of natural hydrogels and will potentially benefit the studies of biomimetic and bioinspired biomaterials.

Atomic Force Microscopy Study of Cross-Sections of Perovskite Layers

Improvement of methods for imaging of the volume structure of photoactive layers is one of the important directions towards development of highly efficient solar cells. In particular, volume structure of photoactive layer has critical influence on perovskite solar cell performance and life time. In this study, a perovskite photoactive layer cross-section was prepared by using Focused Ion Beam (FIB) and imaged by Atomic Force Microscopy (AFM) methods. The proposed approach allows using advances of AFM for imaging structure of perovskites in volume. Two different types of perovskite layers was investigated: FAPbBr3 and MAPbBr3. The heterogeneous structure inside film, which consist of large crystals penetrating the film as well as small particles with sizes of several tens nanometers, is typical for FAPbBr3. The ordered nanocrystalline structure with nanocrystals oriented at 45 degree to film surface is observed in MAPbBr3. An optimized sample preparation route, which includes FIB surface polishing by low energy Ga ions at the angles around 10 degree to surface plane, is described and optimal parameters of surface treatment are discussed. Use of AFM phase contrast method provides high contrast imaging of perovskite structure due to strong dependence of phase shift of oscillating probe on materials properties. The described method of imaging can be used for controllable tuning of perovskite structure by changes of the sample preparation routes.

Nanoscale Multiparametric Imaging of Peptide-Assembled Nanofibrillar Hydrogels by Atomic Force Microscopy

Peptide-assembled hydrogels have emerged as a promising biomaterial for tissue engineering due to their controllable structures as well as biocompatible and biodegradable characteristics. However, the detailed situations of nanofibrillar mechanics during the formation of peptide-assembled hydrogels remain poorly understood. The advent of atomic force microscopy (AFM) provides a powerful tool for nanoscale characterizations and measurements of native biomaterials without pretreatments. In this paper, AFM peak force tapping multiparametric imaging was utilized to investigate the morphology and mechanics of individual nanofibrils during the gelation and degradation process of peptide-assembly hydrogels. Nanofibrillar hydrogels were fabricated by the self-assembly reactions of phenylalanine peptide molecules. AFM topography imaging distinctly visualized the diverse assembly behaviors of peptide-formed nanofibrils during the gelation and degradation of hydrogels, which were significantly correlated with the changes of nanofibrillar mechanics (Young's modulus, adhesion force, deformation, and persistence length) revealed by AFM mechanical imaging. The research provides a novel idea for nanoscale imaging and mechanical analysis of nanofibrillar behaviors in peptide-assembled hydrogels, which will have potential impacts on the characterization and design of supramolecular biomaterials.

Atomic Force Microscopy in Probing Tumor Physics for Nanomedicine

Investigating the physics in tumor development and progression has been recognized as a promising way to advance both basic cancer research and clinical cancer treatment. The advent of atomic force microscopy (AFM) provides an exciting multifunctional tool for investigating the physical properties of biological systems under aqueous conditions with unprecedented spatiotemporal resolution, opening new possibilities for nanomedicine in physical oncology. In this paper, the biomedical applications of AFM to investigate the physical sciences in oncology at single-cell, single-molecule, and tissue levels are systematically reviewed, taking lymphomas as an example. First, detecting the physics of cancers at multiple scales is summarized, along with antibody-based therapy for lymphomas. Next, the principles and methods of AFM imaging and measurements are summarized. After that, utilizing AFM to probe the physics of tumors is reviewed from several facets, including topographical imaging, mechanical measurement, molecular recognition, primary cell detection, and tissue characterization. In particular, the motivations and state-of-the-art overviews of each facet are presented. Finally, the current limitations and future directions of AFM assays for cancer investigations are discussed.

Advances in atomic force microscopy for single-cell analysis

Single-cell analysis has been considered as a promising way to uncover the underlying mechanisms guiding the mysteries of life activities, which considerably complements traditional ensemble assays and yields novel insights into cell biology. The advent of atomic force microscopy (AFM) provides a potent tool for investigating the structures and properties of native biological samples at the micro/nanoscale under near-physiological conditions, which promotes the studies of single-cell behaviors. In the past decades, AFM has achieved great success in single-cell observation and manipulation for biomedical applications, demonstrating the excellent capabilities of AFM in addressing biological issues at the single-cell level with unprecedented spatiotemporal resolution. In this article, we review the recent advances in single-cell analysis that has been made with the utilization of AFM, and provide perspectives for future progression.

A Review of Nanoscale Characterizing Individual DNA Behaviors Using Atomic Force Microscopy

Deoxyribonucleic acid (DNA), the structural basis of genetics, plays a crucial role in the life of cells. The abnormal behaviors of DNA are closely related to the pathological changes of living organisms. The advent of atomic force microscopy (AFM) provides an exciting instrument for biophysically probing the nanoscale dynamic activities of the individual native molecules, considerably complementing the results obtained by traditional biochemical population-averaged measurements. In the past decades, AFM has been widely used in DNA studies, significantly providing novel insights into molecular biology. In this paper, the methodologies and progresses of utilizing AFM to characterize the behaviors of individual DNA molecules from several facets (including DNA topography imaging, DNA mechanics measurements, DNA origami, and DNA nanorobotics) are reviewed, and the future directions are discussed.

The Kinetoplast of Trypanosomatids: From Early Studies of Electron Microscopy to Recent Advances in Atomic Force Microscopy

The kinetoplast is a specialized region of the mitochondria of trypanosomatids that harbors the most complex and unusual mitochondrial DNA found in nature. Kinetoplast DNA (kDNA) is composed of thousands of circular molecules topologically interlocked to form a single network. Two types of DNA circles are present in the kinetoplast: minicircles (0.5–10 kb) and maxicircles (20–40 kb). Knowledge of kinetoplast architecture is crucial to understanding the replication and segregation of kDNA circles because the molecules involved in these processes are precisely positioned in functional domains throughout the kinetoplast. The fine structure of the kinetoplast was revealed in early electron microscopy (EM) studies. However, an understanding of the topological organization of kDNA was only demonstrated after the development of protocols to separate kDNA from nuclear DNA, followed by EM observations. Electron microscopy analysis of thin sections of trypanosomatids, spreading of isolated kDNA networks onto EM grids, deep-etching studies, and cytochemical and immunocytochemical approaches are examples of techniques that were useful for elucidating the structure and replication of the kinetoplast. Recently, atomic force microscopy has joined this set of techniques and improved our knowledge about the kDNA network and revealed new details about kDNA topology in trypanosomatids.

Nanoscale characterization of dynamic cellular viscoelasticity by atomic force microscopy with varying measurement parameters.

Cell mechanics plays an important role in regulating the physiological activities of cells. The advent of atomic force microscopy (AFM) provides a novel powerful instrument for quantifying the mechanics of single cells at the nanoscale. The applications of AFM in single-cell mechanical assays in the past decade have significantly contributed to the field of cell and molecular biology. However, current AFM-based cellular mechanical studies are commonly carried out with fixed measurement parameters, which provides limited information about the dynamic cellular mechanical behaviors in response to the variable external stimuli. In this work, we utilized AFM to investigate cellular viscoelasticity (portrayed as relaxation time) with varying measurement parameters, including ramp rate and surface dwell time, on both cell lines and primary cells. The experimental results show that the obtained cellular relaxation times are remarkably dependent on the parameter surface dwell time and ramp rate during measurements. Besides, the dependencies to the measurement parameters are variable for different types of cells, which can be potentially used to indicate cell states. The research improves our understanding of single-cell dynamic rheology and provides a novel idea for discriminating different types of cells by AFM-based cellular viscoelastic assays with varying measurement parameters.

Advances in Atomic Force Microscopy for Probing Polymer Structure and Properties

Over the past 30 years, atomic force microscopy (AFM) has played an important role in elucidating the structure and properties of polymer surfaces. AFM-based techniques have enabled the quantitative determination of the physicochemical properties of polymer surfaces with high spatial resolution and under a wide variety of conditions. Coupled with the improvements in spatial and temporal resolution, multiparametric and multifunctional characterization has revealed the delicate interplay between structure, dynamics, and properties at the surfaces of complex systems. Here we summarize some of the significant advances that have been made in synthetic polymeric materials, most in the past 10 years, where AFM has been crucial, and we provide our perspective on where AFM will be insightful in future and instrumental in advancing emerging areas.

Atomic force microscopy analysis of extracellular vesicles.

Extracellular vesicles (EVs) are small vesicles ensuring transport of molecules between cells and throughout the body. EVs contain cell type-specific signatures and have been proposed as biomarkers in a variety of diseases. Their small size (<1μm) and biological and physical functions make them obvious candidates for therapeutic agents in immune therapy, vaccination, regenerative medicine and drug delivery. However, due to the complexity and heterogeneity of their origin and composition, the actual mechanism through which these vesicles exert their functions is still unknown and represents a great biomedical challenge. Moreover, because of their small dimensions, the quantification, size distribution and biophysical characterization of these particles are challenging and still subject to controversy. Here, we address the advantage of atomic force microscopy (AFM), for the characterization of isolated EVs. We review AFM imaging of EVs immobilized on different substrates (mica, glass) to identify the influence of isolation and deposition methods on the size distribution, morphology and mechanical properties of EVs.

AFM-Nanomechanical Test: An Interdisciplinary Tool That Links the Understanding of Cartilage and Meniscus Biomechanics, Osteoarthritis Degeneration, and Tissue Engineering.

Our objective is to provide an in-depth review of the recent technical advances of atomic force microscopy (AFM)-based nanomechanical tests and their contribution to a better understanding and diagnosis of osteoarthritis (OA), as well as the repair of tissues undergoing degeneration during OA progression. We first summarize a range of technical approaches for AFM-based nanoindentation, including considerations in both experimental design and data analysis. We then provide a more detailed description of two recently developed modes of AFM-nanoindentation, a high-bandwidth nanorheometer system for studying poroviscoelasticity and an immunofluorescence-guided nanomechanical mapping technique for delineating the pericellular matrix (PCM) and territorial/interterritorial matrix (T/IT-ECM) of surrounding cells in connective tissues. Next, we summarize recent applications of these approaches to three aspects of joint-related healthcare and disease: cartilage aging and OA, developmental biology and OA pathogenesis in murine models, and nanomechanics of the meniscus. These studies were performed over a hierarchy of length scales, from the molecular, cellular to the whole tissue level. The advances described here have contributed greatly to advancing the fundamental knowledge base for improved understanding, detection, and treatment of OA.

Atomic Force Microscopy in Characterizing Cell Mechanics for Biomedical Applications: A Review.

Cell mechanics is a novel label-free biomarker for indicating cell states and pathological changes. The advent of atomic force microscopy (AFM) provides a powerful tool for quantifying the mechanical properties of single living cells in aqueous conditions. The wide use of AFM in characterizing cell mechanics in the past two decades has yielded remarkable novel insights in understanding the development and progression of certain diseases, such as cancer, showing the huge potential of cell mechanics for practical applications in the field of biomedicine. In this paper, we reviewed the utilization of AFM to characterize cell mechanics. First, the principle and method of AFM single-cell mechanical analysis was presented, along with the mechanical responses of cells to representative external stimuli measured by AFM. Next, the unique changes of cell mechanics in two types of physiological processes (stem cell differentiation, cancer metastasis) revealed by AFM were summarized. After that, the molecular mechanisms guiding cell mechanics were analyzed. Finally the challenges and future directions were discussed.

Imaging and Force Recognition of Single Molecular Behaviors Using Atomic Force Microscopy.

The advent of atomic force microscopy (AFM) has provided a powerful tool for investigating the behaviors of single native biological molecules under physiological conditions. AFM can not only image the conformational changes of single biological molecules at work with sub-nanometer resolution, but also sense the specific interactions of individual molecular pair with piconewton force sensitivity. In the past decade, the performance of AFM has been greatly improved, which makes it widely used in biology to address diverse biomedical issues. Characterizing the behaviors of single molecules by AFM provides considerable novel insights into the underlying mechanisms guiding life activities, contributing much to cell and molecular biology. In this article, we review the recent developments of AFM studies in single-molecule assay. The related techniques involved in AFM single-molecule assay were firstly presented, and then the progress in several aspects (including molecular imaging, molecular mechanics, molecular recognition, and molecular activities on cell surface) was summarized. The challenges and future directions were also discussed.

The dynamic interactions between chemotherapy drugs and plasmid DNA investigated by atomic force microscopy

The advent of atomic force microscopy (AFM) provides a powerful tool for imaging individual DNA molecules. Chemotherapy drugs are often related to DNAs. Though many specific drug-DNA interactions have been observed by AFM, knowledge about the dynamic interactions between chemotherapy drugs and plasmid DNAs is still scarce. In this work, AFM was applied to investigate the nanoscale interactions between plasmid DNAs and two commercial chemotherapy drugs (methotrexate and cisplatin). Plasmid DNAs were immobilized on mica which was coated by silanes in advance. AFM imaging distinctly revealed the dynamic changes of single plasmid DNAs after the stimulation of methotrexate and cisplatin. Geometric features of plasmid DNAs were extracted from AFM images and the statistical results showed that the geometric features of plasmid DNAs changed significantly after the stimulation of drugs. This research provides a novel idea to study the actions of chemotherapy drugs against plasmid DNAs at the single-molecule level.

Nanoscale imaging and force probing of biomolecular systems using atomic force microscopy: from single molecules to living cells.

Due to the lack of adequate tools for observation, native molecular behaviors at the nanoscale have been poorly understood. The advent of atomic force microscopy (AFM) provides an exciting instrument for investigating physiological processes on individual living cells with molecular resolution, which attracts the attention of worldwide researchers. In the past few decades, AFM has been widely utilized to investigate molecular activities on diverse biological interfaces, and the performances and functions of AFM have also been continuously improved, greatly improving our understanding of the behaviors of single molecules in action and demonstrating the important role of AFM in addressing biological issues with unprecedented spatiotemporal resolution. In this article, we review the related techniques and recent progress about applying AFM to characterize biomolecular systems in situ from single molecules to living cells. The challenges and future directions are also discussed.