Nano-Mechanical Properties of Modified Poly(Methyl Methacrylate) Films Studied by Atomic Force Microscopy

Atomic force microscopy (AFM) has been used to study the effect of various photoinitiators doped into poly(methyl methacrylate) (PMMA) on the mechanical properties of PMMA films at the nanometer scale. Pure and modified PMMA films (containing four different photoinitiators) were exposed to a mercury vapor lamp in air atmosphere. Force–distance curves for hardness, Young modulus, and adhesion forces were obtained using different AFM modes (tapping or contact-mode) and different tips (diamond or silicon nitride). The results revealed that the added photoinitiators slightly changed the nanomechanical properties of PMMA as a result of alterations in the photochemical reactions and physical processes occurring in the studied systems. tert-Butyl peroxybenzoate had the most efficient effect on the measured parameters in UV-irradiated PMMA, whereas benzoyl peroxide was less active. The mechanism of the observed processes is discussed.     

Investigation of Wear Particles Generated in Human Knee Joints Using Atomic Force Microscopy

Wear particle analysis can be potentially developed as an effective method for assessment of osteoarthritis (OA). To achieve this goal, the surface morphological and mechanical properties of human wear particles extracted from the osteoarthritic synovial joints with different OA grades need to be studied. Atomic force microscopy (AFM) has been used for cartilage analysis owing to its high resolution and the capability of revealing both mechanical properties and surface topographical data in three-dimensions. Few studies have been conducted on human wear particles due to difficulties in obtaining the samples and technical challenges in preparing wear debris samples for AFM investigations in a hydrated environment. This work aimed to develop a suitable preparation technique to study the mechanical properties and surface morphology of human wear particles using AFM. Wear particles were separated from synovial fluid samples which were collected from OA patients and deposited on an aldehyde functional plasma polymer surface to immobilise wear particles. They were imaged for the first time using AFM. The nanoscaled surface topographies and nanomechanical properties of the particles were obtained in a hydrated mode. The methodology established in this study enables investigations of the surface morphology and mechanical properties of wear particles at the nanoscale for better understanding of OA and the possibility of developing a new diagnostic method based on the wear debris analysis technique.     

Indentation modulus and hardness of viscoelastic thin films by atomic force microscopy: A case study

We propose a nanoindentation technique based on atomic force microscopy (AFM) that allows one to deduce both indentation modulus and hardness of viscoelastic materials from the force versus penetration depth dependence, obtained by recording the AFM cantilever deflection as a function of the sample vertical displacement when the tip is pressed against (loading phase) and then removed from (unloading phase) the surface of the sample. Reliable quantitative measurements of both indentation modulus and hardness of the investigated sample are obtained by calibrating the technique through a set of different polymeric samples, used as reference materials, whose mechanical properties have been previously determined by standard indentation tests. By analyzing the dependence of the cantilever deflection versus time, the proposed technique allows one to evaluate and correct the effect of viscoelastic properties of the investigated materials, by adapting a post-experiment data processing procedure well-established for standard depth sensing indentation tests. The technique is described in the case of the measurement of indentation modulus and hardness of a thin film of poly(3,4-ethylenedioxythiophene) doped with poly(4-styrenesulfonate), deposited by chronoamperometry on an indium tin oxide (ITO) substrate.     

In-plane contributions to phase contrast in intermittent contact atomic force microscopy

Contrast in the phase response of intermittent-contact atomic force microscopy (IC-AFM) reveals in-plane structural and mechanical properties of polymer monolayers. This result is unexpected, as IC-AFM has previously only been considered as a probe of out-of-plane properties. Until now, AFM measurements of nanoscale in-plane properties have employed contact mode techniques. In-plane property measurements are possible with intermittent contact AFM because there is a small but significant component of tip motion parallel to the sample surface. This in-plane component of tip displacement is virtually universal in AFM, implying that oscillating-tip techniques generally are sensitive to in-plane material properties. We present a simple Hertzian model of intermittent-contact AFM that includes such an in-plane displacement.     

Mechanical properties of plasma membrane and nuclear envelope measured by scanning probe microscope

Atomic force microscopy has been used to visualize nano‐scale structures of various cellular components and to characterize mechanical properties of biomolecules. In spite of its ability to measure non‐fixed samples in liquid, the application of AFM for living cell manipulation has been hampered by the lack of knowledge of the mechanical properties of living cells. In this study, we successfully combine AFM imaging and force measurement to characterize the mechanical properties of the plasma membrane and the nuclear envelope of living HeLa cells in a culture medium. We examine cantilevers with different physical properties (spring constant, tip angle and length) to find out the one suitable for living cell imaging and manipulation. Our results of elasticity measurement revealed that both the plasma membrane and the nuclear envelope are soft enough to absorb a large deformation by the AFM probe. The penetrations of the plasma membrane and the nuclear envelope were possible when the probe indents the cell membranes far down close to a hard glass surface. These results provide useful information to the development of single‐cell manipulation techniques.     

Substrate influence on cell shape and cell mechanics: HepG2 cells spread on positively charged surfaces

A human hepatoma cell line (HepG2) was cultured on positively and negatively charged polyelectrolytes. Cell/surface adhesion and cell shape evolution were followed with quartz microbalance with dissipation (QCM‐D) and optical microscopy as a function of time, respectively. In particular, substrates coated with poly(ethyleneimine) (PEI) led to fast cell attachment and further spreading, with average maximum frequency Δf = 79 Hz and dissipation ΔD = 40 × 10^?6. On the contrary, no cell spreading was observed on poly(sodium‐4‐styrenesulfonate) (PSS), with Δf = 33 Hz and ΔD = 4.5 × 10^?6. Atomic force microscopy (AFM) was used to investigate the influence of cell shape on its mechanical properties. Considering the cells as an homogenous solid material, the corresponding elastic modulus was estimated using the Hertz model. The elastic modulus was calculated at the central part of the cell, and the average values obtained were 191 ± 14 Pa and 941 ± 58 Pa for cells adsorbed on PSS and PEI, respectively. Thus, different cell–substrate interaction implied different cell mechanical properties reflected in a higher elastic modulus for stronger cell/substrate interaction. The combination of QCM‐D, AFM, and optical microscopy allowed the online study of the cell adhesion process, and the mechanical properties of the adhered cells. Microsc. Res. Tech. 2009. © 2009 Wiley‐Liss, Inc.     

Tribological studies of poly(ether ether ketone) blends

A review of friction and wear studies of poly(ether ether ketone) (PEEK) blended with other polymers is presented. PEEK is a high performance thermoplastic now commonly employed in many engineering applications, but its tribological properties deteriorate in high temperature environments. One approach to improving the friction and wear properties of PEEK is to blend this plastic with appropriate polymers. In recent years, a number of investigations into the tribology of PEEK blended with poly(ether imide) (PEI), polytetrafluoroethylene (PTFE) and liquid crystalline polymers (LCPs) have been reported and these studies are discussed.     

Application of AFM from microbial cell to biofilm

Atomic Force Microscopy (AFM) has proven itself over recent years as an essential tool for the analysis of microbial systems. This article will review how AFM has been used to study microbial systems to provide unique insight into their behavior and relationship with their environment. Immobilization of live cells has enabled AFM imaging and force measurement to provide understanding of the structure and function of numerous microbial cells. At the macromolecular level AFM investigation into the properties of surface macromolecules and the energies associated with their mechanical conformation and functionality has helped unravel the complex interactions of microbial cells. At the level of the whole cell AFM has provided an integrated analysis of how the microbial cell exploits its environment through its selective, adaptable interface, the cell surface. In addition to these areas of study the AFM investigation of microbial biofilms has been vital for industrial and medical process analysis. There exists a tremendous potential for the future application of AFM to microbial systems and this has been strengthened by the trend to use AFM in combination with other characterization methods, such as confocal microscopy and Raman spectroscopy, to elucidate dynamic cellular processes. SCANNING 32: 134–149, 2010. © 2010 Wiley Periodicals, Inc.     

Nanometer-scale heat-conductivity measurements on biological samples

With semiconductor structures reaching the nanometer scale, heat conductivity measurements on the mesoscopic range of some tens of nanometers become an increasingly important aspect for the further improvement in digital processing and storage. Also the attempt to use atomic-force microscopy (AFM) technology for high-density data storage by writing information bits as nanometer-sized indentations into a polymer substrate with a heated cantilever tip asks for a careful investigation of the nano-scale heat-conductivity properties of polymers. Furthermore, in many AFM imaging applications, heat conductivity can provide additional information about the material the imaged structures consist of. In this respect, heat conductivity can also become very interesting in studies of usually quite heterogeneous biological samples, if the resolution can attain the nanometer range. In standard scanning thermal microscopy application, the tip forms a thermocouple, which precludes high-resolution imaging, as thermocouples cannot be made sufficiently small. In this paper, which focuses on biological applications, we demonstrate that by using an ultra sharp AFM cantilever with a Joule heating element above the tip structure different molecular components can be distinguished thanks to their different heat-conductivity properties. In this case, the resolution is determined by the actual tip size, and it can reach 10nm.     

The effect of fluid properties and geometrical parameters of cantilever on the frequency response of atomic force microscopy

Nowadays, the atomic force microscopy plays an indispensable role in imaging and manipulation of biological samples. To observe some specific behaviors and biological processes, fast and accurate imaging techniques are required, and one way to speed up the imaging process is to use short cantilevers. For short beams, the Timoshenko model seems to be more accurate compared to other models such as the Euler–Bernoulli. By using the Timoshenko beam model, the effects of rotational inertia and shear deformation are taken into consideration. In this paper, the frequency response of a rectangular atomic force microscope (AFM) in liquid environment has been analyzed by using the Timoshenko beam model. Afterward, since the dynamic response of AFM is influenced by the applied medium, the effects of physical and mechanical properties (e.g., fluid density and viscosity) on the frequency response of the system have been investigated. The frequency responses of the AFM cantilever immersed in various liquids have been compared with one another. And eventually, to study the influence of geometry on the dynamic behavior of AFM, the effect of the cantilever's geometrical parameters (e.g., cantilever length, width and thickness) on the frequency response of the system has been studied.     

Quantitative measurement of indentation hardness and modulus of compliant materials by atomic force microscopy

An atomic force microscopy (AFM) based technique is proposed for the characterization of both indentation modulus and hardness of compliant materials. A standard AFM tip is used as an indenter to record force versus indentation curves analogous to those obtained in standard indentation tests. In order to overcome the lack of information about the apex geometry, the proposed technique requires calibration using a set of reference samples whose mechanical properties have been previously characterized by means of an independent technique, such as standard indentation. Due to the selected reference samples, the technique has been demonstrated to allow reliable measurements of indentation modulus and hardness in the range of 0.3-4.0 GPa and 15-250 MPa, respectively.     

原子力显微镜在聚合物研究中的应用

原子力显微镜以其分辨率高、样品无需特殊制备、实验可在大气环境中进行等优点而广泛应用于聚合物研究之中,弥补扫描隧道显微镜不能观测非导电样品的缺憾。近年来,其应用已由对聚合物表面几何形貌的观测发展到纳米级结构和表面性能的研究领域。在介绍原子力显微镜工作原理的基础上,简要回顾其在聚合物研究方面的若干新应用,并对其应用前景作展望。     

Measuring biomaterials mechanics with atomic force microscopy. 1. Influence of the loading rate and applied force (pyramidal tips)

Atomic force microscopy (AFM) is today an established tool in imaging and determination of mechanical properties of biomaterials. Due to their complex organization, those materials show intricate properties such as viscoelasticity. Therefore, one has to consider that the loading rate at which the sample is probed will lead to different mechanical response (properties). In this work, we studied the dependence of the mechanical properties of endothelial cells on the loading rate using AFM in force spectroscopy mode. We employed a sharp, four‐sided pyramidal indenter and loading rates ranging from 0.5 to 20 μm/s. In addition, by variation of the load (applied forces from 100 to 10,000 pN), the dependence of the cell properties on indentation depth could be determined. We then showed that the mechanical response of endothelial cells depends nonlinearly on the loading rate and follows a weak power‐law. In addition, regions of different viscous response at varying indentation depth could be determined. Based on the results we obtained, a general route map for AFM users for design of cell mechanics experiments was described.     

Monitoring of glass derivatization with pulsed force mode atomic force microscopy

Non-specific adsorption of proteins at solid/liquid interfaces is a major problem in the use of synthetic biomaterials and in ultrasensitive detection methods. Grafting surfaces with a dense layer of poly(ethylene glycol) (PEG) or other polymers is a most widely used strategy to solve this task. While such modified surfaces have been characterized by their ability to resist protein adsorption, the polymer layers themselves have rarely been studied in fine detail. Atomic force microscopy (AFM) using the pulsed force mode (PFM), is an ideal technique to investigate structural features and physiochemical properties of surfaces because topology and adhesion are simultaneously detected with high lateral resolution. In the present study, PFM-AFM was applied to thoroughly characterize different stages of glass derivatization, up to the formation of a dense PEG layer. Lateral inhomogeneities in topology and/or adhesion were observed at all stages before PEG attachment. The covalently bound PEG, however, was seen to form a densely packed monolayer with maximal thickness, smooth surface, and weak adhesion. Thus, PFM-AFM appears to be a valuable tool for the characterization of protein-repelling surfaces in solution.     

Changes in the biomechanical properties of a single cell induced by nonthermal atmospheric pressure micro‐dielectric barrier discharge plasma

Mechanical properties of a single cell are closely related to the fate and functions of the cell. Changes in mechanical properties may cause diseases or cell apoptosis. Selective cytotoxic effects of nonthermal atmospheric pressure micro‐dielectric barrier discharge (DBD) plasma have been demonstrated on cancer cells. In this work, changes in the mechanical properties of a single cell induced by nonthermal atmospheric pressure micro‐DBD plasma were investigated using atomic force microscopy (AFM). Two cervical cancer cell lines (HeLa and SiHa) and normal human fibroblast cells (HFBs) were exposed to micro‐DBD plasma for various exposure times. The elasticity of a single cell was determined by force–distance curve measurement using AFM. Young's modulus was decreased by plasma treatment for all cells. The Young's modulus of plasma‐treated HeLa cells was decreased by 75% compared to nontreated HeLa cells. In SiHa cells and HFBs, elasticity was decreased slightly. Chemical changes induced by the plasma treatment, which were observed by Raman spectroscopy, were also significant in HeLa cells compared to SiHa cells and HFBs. These results suggested that the molecular changes induced by micro‐DBD plasma were related to cell mechanical changes.     

QD as a bifunctional cell-surface marker for both fluorescence and atomic force microscopy

Fluorescent quantum dots (QDs) are a new class of fluorescent label and have been extensively used in cell imaging. Streptavidin-conjugated QDs have a diameter of ca. 10–15 nm; therefore when used as probes to label cell-surface biomolecules, they can provide contrast enhancement under atomic force microscopy (AFM) and allow specific proteins to be distinguished from the background. In addition, the size and fluorescent properties potentially make them as probes in correlative fluorescence microscopy (FM) and AFM. In this study, we tested the feasibility of using QD-streptavidin conjugates as probes to label wheat germ agglutinin (WGA) receptors on the membrane of human red blood cells (RBCs) and simultaneously obtain fluorescence and AFM images. The results show that the distribution of QDs labeled on human RBCs was non-uniform and that the number of labeled QDs on different erythrocytes varied significantly, which perhaps indicates different ages of the erythrocytes. Thus, QDs may be employed as bifunctional cell-surface markers for both FM and AFM to quantitatively investigate the distribution and expression of membrane proteins or receptors on cell surface.     

Iterative control approach to high-speed force-distance curve measurement using AFM: time-dependent response of PDMS example

Force-distance curve measurements using atomic force microscope (AFM) has been widely used in a broad range of areas. However, currently force-curve measurements are hampered the its low speed of AFM. In this article, a novel inversion-based iterative control technique is proposed to dramatically increase the speed of force-curve measurements. Experimental results are presented to show that by using the proposed control technique, the speed of force-curve measurements can be increased by over 80 times--with no loss of spatial resolution--on a commercial AFM platform and with a standard cantilever. High-speed force curve measurements using this control technique are utilized to quantitatively study the time-dependent elastic modulus of poly(dimethylsiloxane) (PDMS). The force-curves employ a broad spectrum of push-in (load) rates, spanning two-order differences. The elastic modulus measured at low-speed compares well with the value obtained from dynamic mechanical analysis (DMA) test, and the value of the elastic modulus increases as the push-in rate increases, signifying that a faster external deformation rate transitions the viscoelastic response of PDMS from that of a rubbery material toward a glassy one.     

Artesunate induced morphological and mechanical changes of Jurkat cell studied by AFM

In this study, we have used atomic force microscopy (AFM) to study the morphology and mechanical property changes of Jurkat cells exposed to different concentrations of Artesunate (ART) for 24 h at single cellular level. Cell viability and proliferation assays were performed by using the Cell Counting Kit‐8. The concentration of ART, which resulted in the inhibition rate >50% was selected. The AFM images revealed that the cell membrane changed and the ultrastructure also became complex. Mechanical properties of individual cell were tracked with AFM‐based force spectroscopy. The force curves revealed that when a cell was exposed to the ART, the mechanical properties changed obviously. Treated cells had a lower adhesion force of 416.8±37.9 pN, whereas control group had a higher adhesion force of 1064.2±97.0 pN. The Young's modulus decreased to nearly one‐third, from control group of 0.648±0.037 kPa to treated group of 0.254±0.035 kPa and the stiffness increased to nearly 1.5 times, from control group of 1.231±0.084 mN/m to treated group of 1.917±0.137 mN/m. These results suggest that ART can inhibit the proliferation of Jurkat and induce changes in the morphological structure and mechanical properties of Jurkat cells. The high resolution and high sensitivity of AFM can be used to detect morphological and mechanical properties of cells exposed to ART. The AFM may be developed to be a useful tool for detecting the cell death and evaluating the anti‐carcinogen efficacy against tumor cell. SCANNING 31: 83–89, 2009. © 2009 Wiley Periodicals, Inc.     

Scratching maps for polymers

The scratching technique has gained interest in recent times due to its varied applications to a number of engineering materials, especially for the evaluation of surface scratch resistance of plastics. Scratching provides a convenient and reliable means to investigate the mechanical properties of organic polymers under various contact conditions. The scratch hardness method is widely adopted to provide a first-order evaluation of the relative scratch resistance of materials for comparison purposes. The method also allows the identification and the assessment of the surface deformation processes and maps defining the scratch deformation modes as a function of contact conditions may be generated. These scratching maps may present experimental results in terms of the deformation mechanism, the scratch hardness and the friction coefficient. This paper primarily provides a review of the application of scratching maps for polymers. Results for the scratch hardness and the deformation mechanisms for a poly(methylmethacrylate) (PMMA), a poly(tetrafluoroethylene) (PTFE) and an ultra-high molecular weight poly(ethylene) (UHMWPE) are presented. The PTFE system is also described following the effects of γ-irradiation; radiation produces a marked reduction in toughness. The scratches were produced on the polymer surfaces by cones and spheres of various size under a number of contact conditions (e,g, applied normal load, strain, scratch velocity, etc.). SEM imaging and laser profilometry are used for the study of the deformation mechanisms and the measurements of the scratch profiles. It is shown that polymers exhibit a wide range of scratch deformation characteristics and that the deformation mechanism is determined by the most efficient energy dissipation process for the particular external constraints.     

Assembling and imaging of his-tag green fluorescent protein on mica surfaces studied by atomic force microscopy and fluorescence microscopy

The adsorption of his-tag green fluorescent protein (GFPH(6)) onto the mica surfaces has been studied by atomic force microscopy (AFM) and laser confocal fluorescence microscopy. By controlling the adsorption conditions, separated single GFPH(6) and GFPH(6) monolayer can be adsorbed and formed on mica surfaces. In present experiments, based on the AFM measurement, we found that the adsorbed GFPH(6) was bound on the mica surface with its beta-sheets. The formed GFPH(6) monolayer on mica surfaces was flat, uniform, and stable. Some applications of the formed monolayer have been demonstrated. The formed monolayer can be used as a substrate for DNA imaging and AFM mechanical lithography.