Cell Membrane Oscillations under Radiofrequency Electromagnetic Modulation.

Cell responses to external radiofrequencies (RF) are a fundamental problem of much scientific research, clinical applications, and even daily lives surrounded by wireless communication hardware. In this work, we report an unexpected observation that the cell membrane can oscillate at the nanometer scale in phase with the external RF radiation from kHz to GHz. By analyzing the oscillation modes, we reveal the mechanism behind the membrane oscillation resonance, membrane blebbing, the resulting cell death, and the selectivity of plasma-based cancer treatment based on the difference in the membrane's natural frequencies among cell lines. Therefore, a selectivity of treatment can be achieved by aiming at the natural frequency of the target cell line to focus the membrane damage on the cancer cells and avoid normal tissues nearby. This gives a promising cancer therapy that is especially effective in the mixing lesion of the cancer cells and normal cells such as glioblastoma where surgical removal is not applicable. Along with these new phenomena, this work provides a general understanding of the cell coupling with RF radiation from the externally stimulated membrane behavior to the cell apoptosis and necrosis.

Optimizing the accuracy of viscoelastic characterization with AFM force-distance experiments in the time and frequency domains.

Atomic Force Microscopy (AFM) force-distance (FD) experiments have emerged as an attractive alternative to traditional micro-rheology measurement techniques owing to their versatility of use in materials of a wide range of mechanical properties. Here, we show that the range of time dependent behaviour which can reliably be resolved from the typical method of FD inversion (fitting constitutive FD relations to FD data) is inherently restricted by the experimental parameters: sampling frequency, experiment length, and strain rate. Specifically, we demonstrate that violating these restrictions can result in errors in the values of the parameters of the complex modulus. In the case of complex materials, such as cells, whose behaviour is not specifically understood a priori, the physical sensibility of these parameters cannot be assessed and may lead to falsely attributing a physical phenomenon to an artifact of the violation of these restrictions. We use arguments from information theory to understand the nature of these inconsistencies as well as devise limits on the range of mechanical parameters which can be reliably obtained from FD experiments. The results further demonstrate that the nature of these restrictions depends on the domain (time or frequency) used in the inversion process, with the time domain being far more restrictive than the frequency domain. Finally, we demonstrate how to use these restrictions to better design FD experiments to target specific timescales of a material's behaviour through our analysis of a polydimethylsiloxane (PDMS) polymer sample.

Frequency-dependent nanomechanical profiling for medical diagnosis.

Atomic force microscopy (AFM), developed in the early 1980s, has become a powerful characterization tool in micro- and nanoscale science. In the early 1990s, its relevance within biology and medicine research became evident, although its incorporation into healthcare applications remains relatively limited. Here, we briefly explore the reasons for this low level of technological adoption. We also propose a path forward for the incorporation of frequency-dependent nanomechanical measurements into integrated healthcare strategies that link routine AFM measurements with computer analysis, real-time communication with healthcare providers, and medical databases. This approach would be appropriate for diseases such as cancer, lupus, arteriosclerosis and arthritis, among others, which bring about significant mechanical changes in the affected tissues.

Viscoelastic parameterization of human skin cells characterize material behavior at multiple timescales.

Countless biophysical studies have sought distinct markers in the cellular mechanical response that could be linked to morphogenesis, homeostasis, and disease. Here, an iterative-fitting methodology visualizes the time-dependent viscoelastic behavior of human skin cells under physiologically relevant conditions. Past investigations often involved parameterizing elastic relationships and assuming purely Hertzian contact mechanics, which fails to properly account for the rich temporal information available. We demonstrate the performance superiority of the proposed iterative viscoelastic characterization method over standard open-search approaches. Our viscoelastic measurements revealed that 2D adherent metastatic melanoma cells exhibit reduced elasticity compared to their normal counterparts-melanocytes and fibroblasts, and are significantly less viscous than fibroblasts over timescales spanning three orders of magnitude. The measured loss angle indicates clear differential viscoelastic responses across multiple timescales between the measured cells. This method provides insight into the complex viscoelastic behavior of metastatic melanoma cells relevant to better understanding cancer metastasis and aggression.

A new method for obtaining model-free viscoelastic material properties from atomic force microscopy experiments using discrete integral transform techniques.

Viscoelastic characterization of materials at the micro- and the nanoscale is commonly performed with the aid of force-distance relationships acquired using atomic force microscopy (AFM). The general strategy for existing methods is to fit the observed material behavior to specific viscoelastic models, such as generalized viscoelastic models or power-law rheology models, among others. Here we propose a new method to invert and obtain the viscoelastic properties of a material through the use of the Z-transform, without using a model. We present the rheological viscoelastic relations in their classical derivation and their z-domain correspondence. We illustrate the proposed technique on a model experiment involving a traditional ramp-shaped force-distance AFM curve, demonstrating good agreement between the viscoelastic characteristics extracted from the simulated experiment and the theoretical expectations. We also provide a path for calculating standard viscoelastic responses from the extracted material characteristics. The new technique based on the Z-transform is complementary to previous model-based viscoelastic analyses and can be advantageous with respect to Fourier techniques due to its generality. Additionally, it can handle the unbounded inputs traditionally used to acquire force-distance relationships in AFM, such as ramp functions, in which the cantilever position is displaced linearly with time for a finite period of time.

Corrosion Resistance of Sulfur-Selenium Alloy Coatings.

Despite decades of research, metallic corrosion remains a long-standing challenge in many engineering applications. Specifically, designing a material that can resist corrosion both in abiotic as well as biotic environments remains elusive. Here a lightweight sulfur-selenium (S-Se) alloy is designed with high stiffness and ductility that can serve as an excellent corrosion-resistant coating with protection efficiency of ≈99.9% for steel in a wide range of diverse environments. S-Se coated mild steel shows a corrosion rate that is 6-7 orders of magnitude lower than bare metal in abiotic (simulated seawater and sodium sulfate solution) and biotic (sulfate-reducing bacterial medium) environments. The coating is strongly adhesive, mechanically robust, and demonstrates excellent damage/deformation recovery properties, which provide the added advantage of significantly reducing the probability of a defect being generated and sustained in the coating, thus improving its longevity. The high corrosion resistance of the alloy is attributed in diverse environments to its semicrystalline, nonporous, antimicrobial, and viscoelastic nature with superior mechanical performance, enabling it to successfully block a variety of diffusing species.

Correction: Extracting viscoelastic material parameters using an atomic force microscope and static force spectroscopy.

[This corrects the article DOI: 10.3762/bjnano.11.77.].

On the frequency dependence of viscoelastic material characterization with intermittent-contact dynamic atomic force microscopy: avoiding mischaracterization across large frequency ranges.

Atomic force microscopy (AFM) is a widely use technique to acquire topographical, mechanical, or electromagnetic properties of surfaces, as well as to induce surface modifications at the micrometer and nanometer scale. Viscoelastic materials, examples of which include many polymers and biological materials, are an important class of systems, the mechanical response of which depends on the rate of application of the stresses imparted by the AFM tip. The mechanical response of these materials thus depends strongly on the frequency at which the characterization is performed, so much so that important aspects of behavior may be missed if one chooses an arbitrary characterization frequency regardless of the materials properties. In this paper we present a linear viscoelastic analysis of intermittent-contact, nearly resonant dynamic AFM characterization of such materials, considering the possibility of multiple characteristic times. We describe some of the intricacies observed in their mechanical response and alert the reader about situations where mischaracterization may occur as a result of probing the material at frequency ranges or with probes that preclude observation of its viscoelastic behavior. While we do not offer a solution to the formidable problem of inverting the frequency-dependent viscoelastic behavior of a material from dynamic AFM observables, we suggest that a partial solution is offered by recently developed quasi-static force-distance characterization techniques, which incorporate viscoelastic models with multiple characteristic times and can help inform dynamic AFM characterization.

Extracting viscoelastic material parameters using an atomic force microscope and static force spectroscopy.

Atomic force microscopy (AFM) techniques have provided and continue to provide increasingly important insights into surface morphology, mechanics, and other critical material characteristics at the nanoscale. One attractive implementation involves extracting meaningful material properties, which demands physically accurate models specifically designed for AFM experimentation and simulation. The AFM community has pursued the precise quantification and extraction of rate-dependent material properties, in particular, for a significant period of time, attempting to describe the standard viscoelastic response of materials. AFM static force spectroscopy (SFS) is one approach commonly used in pursuit of this goal. It is capable of acquiring rich temporal insight into the behavior of a sample. During AFM-SFS experiments the cantilever base approaches samples with a nearly constant velocity, which is manipulated to investigate different timescales of the mechanical response. This manuscript seeks to build upon our previous work and presents an approach to extracting useful linear viscoelastic information from AFM-SFS experiments. In addition, the basis for selecting and restricting the model parameters for fitting is discussed from the perspective of applying this technique on a practical level. This work begins with a guided discussion that develops a fit function from fundamental laws, continues with conditioning a raw SFS experimental dataset, and concludes with the fit and prediction of viscoelastic response parameters such as storage modulus, loss modulus, loss angle, and compliance. These steps constitute a complete guide to leveraging AFM-SFS data to estimate key material parameters, with a series of detailed insights into both the methodology and supporting analytical choices.

Current measurements in the intermittent-contact mode of atomic force microscopy using the Fourier method: a feasibility analysis.

Atomic force microscopy (AFM) is an important tool for measuring a variety of nanoscale surface properties, such as topography, viscoelasticity, electrical potential and conductivity. Some of these properties are measured using contact methods (static contact or intermittent contact), while others are measured using noncontact methods. Some properties can be measured using different approaches. Conductivity, in particular, is mapped using the contact-mode method. However, this modality can be destructive to delicate samples, since it involves continuously dragging the cantilever tip on the surface during the raster scan, while a constant tip-sample force is applied. In this paper we discuss a possible approach to develop an intermittent-contact conductive AFM mode based on Fourier analysis, whereby the measured current response consists of higher harmonics of the cantilever oscillation frequency. Such an approach may enable the characterization of soft samples with less damage than contact-mode imaging. To explore its feasibility, we derive the analytical form of the tip-sample current that would be obtained for attractive (noncontact) and repulsive (intermittent-contact) dynamic AFM characterization, and compare it with results obtained from numerical simulations. Although significant instrumentation challenges are anticipated, the modelling results are promising and suggest that Fourier-based higher-harmonics current measurement may enable the development of a reliable intermittent-contact conductive AFM method.

Few-cycle Regime Atomic Force Microscopy.

Traditionally, dynamic atomic force microscopy (AFM) techniques are based on the analysis of the quasi-steady state response of the cantilever deflection in terms of Fourier analysis. Here we describe a technique that instead exploits the often disregarded transient response of the cantilever through a relatively modern mathematical tool, which has caused important developments in several scientific fields but that is still quite unknown in the AFM context: the wavelet analysis. This tool allows us to localize the time-varying spectral composition of the initial oscillations of the cantilever deflection when an impulsive excitation is given (as in the band excitation method), a mode that we call the few-cycle regime. We show that this regime encodes very meaningful information about the tip-sample interaction in a unique and extremely sensitive manner. We exploit this high sensitivity to gain detailed insight into multiple physical parameters that perturb the dynamics of the AFM probe, such as the tip radius, Hamaker constant, sample's elastic modulus and height of an adsorbed water layer. We validate these findings with experimental evidence and computational simulations and show a feasible path towards the simultaneous retrieval of multiple physical parameters.

Acquisition of time-frequency localized mechanical properties of biofilms and single cells with high spatial resolution.

Biofilms are a cluster of bacteria embedded in extracellular polymeric substances (EPS) that contain a complex composition of polysaccharides, proteins, and extracellular DNA (eDNA). Desirable mechanical properties of the biofilms are critical for their survival, propagation, and dispersal, and the response of mechanical properties to different treatment conditions also sheds light on biofilm control and eradication in vivo and on engineering surfaces. However, it is challenging yet important to investigate mechanical behaviors of biofilms with a high spatial resolution because biofilms are very heterogeneous. Moreover, biofilms are viscoelastic, and their time-dependent mechanical behavior is difficult to capture. Herein, we develop a powerful technique that combines the high spatial resolution of an atomic force microscope (AFM) with a rigorous history-dependent viscoelastic analysis to deliver highly spatial-localized biofilm properties within a wide time-frequency window. By exploiting the use of static force spectroscopy in combination with an appropriate viscoelastic framework, we highlight the intensive amount of time-dependent information experimentally available that has been largely overlooked. It is shown that this technique provides a detailed nanorheological signature of the biofilms even at the single-cell level. We share the computational routines that would allow any user to perform the analysis from experimental raw data. The detailed localization of mechanical properties in space and in time-frequency domain provides insights into the understanding of biofilm stability, cohesiveness, dispersal, and control.

Visible-Light-Responsive Photocatalyst of Graphitic Carbon Nitride for Pathogenic Biofilm Control.

Pathogenic biofilms raise significant health and economic concerns, because these bacteria are persistent and can lead to long-term infections in vivo and surface contamination in healthcare and industrial facilities or devices. Compared with conventional antimicrobial strategies, photocatalysis holds promise for biofilm control because of its broad-spectrum effectiveness under ambient conditions, low cost, easy operation, and reduced maintenance. In this study, we investigated the performance and mechanism of Staphylococcus epidermidis biofilm control and eradication on the surface of an innovative photocatalyst, graphitic carbon nitride (g-C3N4), under visible-light irradiation, which overcame the need for ultraviolet light for many current photocatalysts (e.g., titanium dioxide (TiO2)). Optical coherence tomography and confocal laser scanning microscopy (CLSM) suggested that g-C3N4 coupons inhibited biofilm development and eradicated mature biofilms under the irradiation of white light-emitting diodes. Biofilm inactivation was observed occurring from the surface toward the center of the biofilms, suggesting that the diffusion of reactive species into the biofilms played a key role. By taking advantage of scanning electron microscopy, CLSM, and atomic force microscopy for biofilm morphology, composition, and mechanical property characterization, we demonstrated that photocatalysis destroyed the integrated and cohesive structure of biofilms and facilitated biofilm eradication by removing the extracellular polymeric substances. Moreover, reactive oxygen species generated during g-C3N4 photocatalysis were quantified via reactions with radical probes and 1O2 was believed to be responsible for biofilm control and removal. Our work highlights the promise of using g-C3N4 for a broad range of antimicrobial applications, especially for the eradication of persistent biofilms under visible-light irradiation, including photodynamic therapy, environmental remediation, food-industry applications, and self-cleaning surface development.

Artifacts in time-resolved Kelvin probe force microscopy.

Kelvin probe force microscopy (KPFM) has been used for the characterization of metals, insulators, and semiconducting materials on the nanometer scale. Especially in semiconductors, the charge dynamics are of high interest. Recently, several techniques for time-resolved measurements with time resolution down to picoseconds have been developed, many times using a modulated excitation signal, e.g., light modulation or bias modulation that induces changes in the charge carrier distribution. For fast modulation frequencies, the KPFM controller measures an average surface potential, which contains information about the involved charge carrier dynamics. Here, we show that such measurements are prone to artifacts due to frequency mixing, by performing numerical dynamics simulations of the cantilever oscillation in KPFM subjected to a bias-modulated signal. For square bias pulses, the resulting time-dependent electrostatic forces are very complex and result in intricate mixing of frequencies that may, in some cases, have a component at the detection frequency, leading to falsified KPFM measurements. Additionally, we performed fast Fourier transform (FFT) analyses that match the results of the numerical dynamics simulations. Small differences are observed that can be attributed to transients and higher-order Fourier components, as a consequence of the intricate nature of the cantilever driving forces. These results are corroborated by experimental measurements on a model system. In the experimental case, additional artifacts are observed due to constructive or destructive interference of the bias modulation with the cantilever oscillation. Also, in the case of light modulation, we demonstrate artifacts due to unwanted illumination of the photodetector of the beam deflection detection system. Finally, guidelines for avoiding such artifacts are given.

Theory of Single-Impact Atomic Force Spectroscopy in liquids with material contrast.

Scanning probe microscopy has enabled nanoscale mapping of mechanical properties in important technological materials, such as tissues, biomaterials, polymers, nanointerfaces of composite materials, to name only a few. To improve and widen the measurement of nanoscale mechanical properties, a number of methods have been proposed to overcome the widely used force-displacement mode, that is inherently slow and limited to a quasi-static regime, mainly using multiple sinusoidal excitations of the sample base or of the cantilever. Here, a different approach is put forward. It exploits the unique capabilities of the wavelet transform analysis to harness the information encoded in a short duration spectroscopy experiment. It is based on an impulsive excitation of the cantilever and a single impact of the tip with the sample. It performs well in highly damped environments, which are often seen as problematic in other standard dynamic methods. Our results are very promising in terms of viscoelastic property discrimination. Their potential is oriented (but not limited) to samples that demand imaging in liquid native environments and also to highly vulnerable samples whose compositional mapping cannot be obtained through standard tapping imaging techniques.

Imaging of viscoelastic soft matter with small indentation using higher eigenmodes in single-eigenmode amplitude-modulation atomic force microscopy.

In this short paper we explore the use of higher eigenmodes in single-eigenmode amplitude-modulation atomic force microscopy (AFM) for the small-indentation imaging of soft viscoelastic materials. In viscoelastic materials, whose response depends on the deformation rate, the tip-sample forces generated as a result of sample deformation increase as the tip velocity increases. Since the eigenfrequencies in a cantilever increase with eigenmode order, and since higher oscillation frequencies lead to higher tip velocities for a given amplitude (in viscoelastic materials), the sample indentation can in some cases be reduced by using higher eigenmodes of the cantilever. This effect competes with the lower sensitivity of higher eigenmodes, due to their larger force constant, which for elastic materials leads to greater indentation for similar amplitudes, compared with lower eigenmodes. We offer a short theoretical discussion of the key underlying concepts, along with numerical simulations and experiments to illustrate a simple recipe for imaging soft viscoelastic matter with reduced indentation.

High-stress study of bioinspired multifunctional PEDOT:PSS/nanoclay nanocomposites using AFM, SEM and numerical simulation.

Bioinspired design has been central in the development of hierarchical nanocomposites. Particularly, the nacre-mimetic brick-and-mortar structure has shown excellent mechanical properties, as well as gas-barrier properties and optical transparency. Along with these intrinsic properties, the layered structure has also been utilized in sensing devices. Here we extend the multifunctionality of nacre-mimetics by designing an optically transparent and electron conductive coating based on PEDOT:PSS and nanoclays Laponite RD and Cloisite Na+. We carry out extensive characterization of the nanocomposite using transmittance spectra (transparency), conductive atomic force microscopy (conductivity), contact-resonance force microscopy (mechanical properties), and SEM combined with a variety of stress-strain AFM experiments and AFM numerical simulations (internal structure). We further study the nanoclay's response to the application of pressure with multifrequency AFM and conductive AFM, whereby increases and decreases in conductivity can occur for the Laponite RD composites. We offer a possible mechanism to explain the changes in conductivity by modeling the coating as a 1-dimensional multibarrier potential for electron transport, and show that conductivity can change when the separation between the barriers changes under the application of pressure, and that the direction of the change depends on the energy of the electrons. We did not observe changes in conductivity under the application of pressure with AFM for the Cloisite Na+ nanocomposite, which has a large platelet size compared with the AFM probe diameter. No pressure-induced changes in conductivity were observed in the clay-free polymer either.

Material property analytical relations for the case of an AFM probe tapping a viscoelastic surface containing multiple characteristic times.

We explore the contact problem of a flat-end indenter penetrating intermittently a generalized viscoelastic surface, containing multiple characteristic times. This problem is especially relevant for nanoprobing of viscoelastic surfaces with the highly popular tapping-mode AFM imaging technique. By focusing on the material perspective and employing a rigorous rheological approach, we deliver analytical closed-form solutions that provide physical insight into the viscoelastic sources of repulsive forces, tip-sample dissipation and virial of the interaction. We also offer a systematic comparison to the well-established standard harmonic excitation, which is the case relevant for dynamic mechanical analysis (DMA) and for AFM techniques where tip-sample sinusoidal interaction is permanent. This comparison highlights the substantial complexity added by the intermittent-contact nature of the interaction, which precludes the derivation of straightforward equations as is the case for the well-known harmonic excitations. The derivations offered have been thoroughly validated through numerical simulations. Despite the complexities inherent to the intermittent-contact nature of the technique, the analytical findings highlight the potential feasibility of extracting meaningful viscoelastic properties with this imaging method.

Analysis and modification of defective surface aggregates on PCDTBT:PCBM solar cell blends using combined Kelvin probe, conductive and bimodal atomic force microscopy.

Organic photovoltaic systems comprising donor polymers and acceptor fullerene derivatives are attractive for inexpensive energy harvesting. Extensive research on polymer solar cells has provided insight into the factors governing device-level efficiency and stability. However, the detailed investigation of nanoscale structures is still challenging. Here we demonstrate the analysis and modification of unidentified surface aggregates. The aggregates are characterized electrically by Kelvin probe force microscopy and conductive atomic force microscopy (C-AFM), whereby the correlation between local electrical potential and current confirms a defective charge transport. Bimodal AFM modification confirms that the aggregates exist on top of the solar cell structure, and is used to remove them and to reveal the underlying active layer. The systematic analysis of the surface aggregates suggests that the structure consists of PCBM molecules.

Imaging of surface nanobubbles by atomic force microscopy in liquids: Influence of drive frequency on the characterization of ultrasoft matter.

Imaging of soft matter with atomic force microscopy (AFM) is challenging due to tip-induced deformation, which convolutes with the measurement. The challenges are generally more serious in liquid environments due to a severe loss of sensitivity of the vibrating microcantilever to external forces, as well as due to the presence of undesirable mechanical resonances when piezoelectric excitation systems are used. Furthermore, the choice of imaging parameters can have a significant impact on the quality of the results, such that the customary practices used for tuning the cantilever are not always appropriate. Here we explore the influence of the chosen drive frequency on the imaging of ultrasoft matter, using surface gas nanobubbles on gold-coated glass and highly oriented pyrolytic graphite (HOPG) as a test platform. We carry out single- and multifrequency AFM experiments using both the traditional amplitude-peak method and the recently proposed phase-slope-peak method for tuning the cantilever, as well as piezoelectric and photothermal excitation, providing an extensive discussion on the factors governing the level of tip-induced sample deformation and the quality of the phase contrast obtained for each of the methods. The general conclusion is that there is no "one-size-fits-all" approach for tuning the cantilever for low-impact tapping-mode AFM, although rational optimization of the imaging process is generally possible, whereby the choice of the drive frequency plays a prominent role. The physical insight and guidelines provided here can be extremely useful for the gentle imaging of a wide range of biological and other soft materials in liquid environments. Microsc. Res. Tech. 80:41-49, 2017. © 2016 Wiley Periodicals, Inc.