AFM Nanoindentation Research Articles

Structural and Mechanical Characterization of DNA-Tethered Membranes on Graphene.

Lipid membranes that are separated from the surface of graphene by DNA tethers were prepared by surface functionalization with pyrene coupled to single-stranded DNA (ssDNA), followed by self-assembly of the mixture of ssDNA-functionalized phospholipid and the matrix phospholipids. The formation of uniform membranes was confirmed by fluorescence microscopy, and the structures of the systems before and after hybridization in the direction perpendicular to the global plane of the membranes were investigated using high-energy X-ray reflectivity. The thickness values of the DNA spacers (15 and 37 bp) calculated from the best-fit results were less than the expected thicknesses of the double-stranded DNA (dsDNA) chains taking the upright conformation, indicating that the DNA spacers are tilted with respect to the direction normal to the surface. The Young's moduli of the DNA-tethered membranes obtained by AFM nanoindentation showed higher values than the membranes with no DNA tethers, which suggests that the DNA layer resists against the compression, lifting up the membrane. Intriguingly, the presence of DNA tethers caused no increase in the yield depth. The smaller thickness values as well as the unchanged yield depth suggest that the dsDNA chains can tilt and rotate, which can be attributed to the flexible pyrene-DNA junction.

In-silico simulation of nanoindentation on bone using a 2D cohesive finite element model

This study proposed and validated a 2D finite element (FE) model for conducting in-silico simulations of in-situ nanoindentation tests on mineralized collagen fibrils (MCF) and the extrafibrillar matrix (EFM) within human cortical bone. Initially, a multiscale cohesive FE model was developed by adapting a previous model of bone lamellae, encompassing both MCF and EFM. Subsequently, nanoindentation tests were simulated in-silico using this model, and the resulting predictions were compared to AFM nanoindentation test data to verify the model's accuracy. The FE model accurately predicted nanoindentation results under wet conditions, closely aligning with outcomes obtained from AFM nanoindentation tests. Specifically, it successfully mirrored the traction/separation curve, nanoindentation modulus, plastic energy dissipation, and plastic energy ratio obtained from AFM nanoindentation tests. Additionally, this in-silico model demonstrated its ability to capture alterations in nanoindentation properties caused by the removal of bound water, by considering corresponding changes in mechanical properties of the collagen phase and the interfaces among bone constituents. Notably, significant changes in the elastic modulus and plastic energy dissipation were observed in both MCF and EFM compartments of bone, consistent with observations in AFM nanoindentation tests. These findings indicate that the proposed in-silico model effectively captures the influence of ultrastructural changes on bone's mechanical properties at sub-lamellar levels. Presently, no experimental methods exist to conduct parametric studies elucidating the ultrastructural origins of bone tissue fragility. The introduction of this in-silico model presents an invaluable tool to bridge this knowledge gap in the future.

Sensitivity in nanomechanical pedestal MEMS cantilever

Nanomechanical resonator-based sensing devices are used in medical diagnostics based on their high-frequency dynamic behavior. Cantilevers fall into the category of Nanomechanical resonators. It also resembles a resonator whose shape is like that of a nanowire clamped at one end. As the surface-to-volume ratio of a nanowire resonator increases due to scaling down, surface stress plays a crucial role in the mechanical behavior of a resonator. Piezoresistive MEMS cantilevers are used for vapor phase analysis of volatile compounds and gas. Studies were done to address the mass sensitivity issues and fractures associated with bioceramic and nanocomposite coatings-based cantilever resonators. The studies show how the sensing performance can be determined or tuned. Finite Element modelling of a pedestal cantilever resonator was done which takes into consideration the mass sensitivity issues associated with linear cantilevers. MEMS cantilever structures were fabricated using lithographic techniques. SiCN, a suitable material for MEMS in harsh operating conditions were deposited on Silicon substrates showing high hardness (20 GPa) and modulus (220 GPa). The electrical conduction taking place between the nanoindentation tip and sample was explored. The surface stress of 711 MPa was determined based on pedestal cantilever deflections, Fracture taking place on the sample surface was found to affect the conduction and tip displacements in AFM nanoindentations. The sharpness of the tip was found to have an influence on the tip-sample conduction mechanism useful for MEMS applications. The sensitivity associated with cell mass detection was reported to get affected by film deposition stress and formation of surface cracks and puts a challenge for doing future work.

Crackling noise microscopy

Crackling noise is a scale-invariant phenomenon found in various driven nonlinear dynamical material systems as a response to external stimuli such as force or external fields. Jerky material movements in the form of avalanches can span many orders of magnitude in size and follow universal scaling rules described by power laws. The concept was originally studied as Barkhausen noise in magnetic materials and now is used in diverse fields from earthquake research and building materials monitoring to fundamental research involving phase transitions and neural networks. Here, we demonstrate a method for nanoscale crackling noise measurements based on AFM nanoindentation, where the AFM probe can be used to study the crackling of individual nanoscale features, a technique we call crackling noise microscopy. The method is successfully applied to investigate the crackling of individual topological defects, i.e. ferroelectric domain walls. We show that critical exponents for avalanches are altered at these nanoscale features, leading to a suppression of mixed-criticality, which is otherwise present in domains. The presented concept opens the possibility of investigating the crackling of individual nanoscale features in a wide range of material systems.

Elasticity and material anisotropy of lamellar cortical bone in adult bovine tibia characterized via AFM nanoindentation

The research focuses on the evaluation of the mechanical properties of osteonal cortical bone at the lamellar level. Elastic properties of the mid-diaphysis region of the bovine tibia are investigated via cantilever-based nanoindentation at the submicron length scale utilizing Atomic Force Microscopy, where the force-displacement curves are used for the elastic assessment using the Derjaguin-Muller-Toropov model to calculate indentation modulus. Variations of the modulus and the directional mechanical response of the osteonal bone at different distances from the Haversian canal are investigated. Additionally, the effects of demineralization on the indentation modulus are discussed. It was found that in the axial direction, the first and last untreated thick lamella layers show a significant indentation modulus difference compared to all other layers (4.26 ± 0.4 and 4.6 ± 0.3 GPa vs ∼3.5 GPa). On the other hand, the indentation modulus of transverse thick lamella layers shows a periodic variation between ∼3 ± 0.7 GPa and ∼4 ± 0.3 GPa from near the Haversian canal to near the interstitial bone. A periodic variation in the anisotropy ratio was found. Mineral content was quantified via energy-dispersive X-ray microanalysis at different levels of mineralization and shows a positive correlation with the indentation modulus.

Three-Dimensional Collagen Topology Shapes Cell Morphology, beyond Stiffness.

Cellular heterogeneity is associated with many physiological processes, including pathological ones, such as morphogenesis and tumorigenesis. The extracellular matrix (ECM) is a key player in the generation of cellular heterogeneity. Advances in our understanding rely on our ability to provide relevant in vitro models. This requires obtainment of the characteristics of the tissues that are essential for controlling cell fate. To do this, we must consider the diversity of tissues, the diversity of physiological contexts, and the constant remodeling of the ECM along these processes. To this aim, we have fabricated a library of ECM models for reproducing the scaffold of connective tissues and the basement membrane by using different biofabrication routes based on the electrospinning and drop casting of biopolymers from the ECM. Using a combination of electron microscopy, multiphoton imaging, and AFM nanoindentation, we show that we can vary independently protein composition, topology, and stiffness of ECM models. This in turns allows one to generate the in vivo complexity of the phenotypic landscape of ovarian cancer cells. We show that, while this phenotypic shift cannot be directly correlated with a unique ECM feature, the three-dimensional collagen fibril topology patterns cell shape, beyond protein composition and stiffness of the ECM. On this line, this work is a further step toward the development of ECM models recapitulating the constantly remodeled environment that cells face and thus provides new insights for cancer model engineering and drug testing.

Stimulus-Responsive, Gelatin-Containing Supramolecular Nanofibers as Switchable 3D Microenvironments for Cells.

Polymer- and/or protein-based nanofibers that promote stable cell adhesion have drawn increasing attention as well-defined models of the extracellular matrix. In this study, we fabricated two classes of stimulus-responsive fibers containing gelatin and supramolecular crosslinks to emulate the dynamic cellular microenvironment in vivo. Gelatin enabled cells to adhere without additional surface functionalization, while supramolecular crosslinks allowed for the reversible switching of the Young’s modulus through changes in the concentration of guest molecules in culture media. The first class of nanofibers was prepared by coupling the host–guest inclusion complex to gelatin before electrospinning (pre-conjugation), while the second class of nanofibers was fabricated by coupling gelatin to polyacrylamide functionalized with host or guest moieties, followed by conjugation in the electrospinning solution (post-conjugation). In situ AFM nano-indentation demonstrated the reversible switching of the Young’s modulus between 2–3 kPa and 0.2–0.3 kPa under physiological conditions by adding/removing soluble guest molecules. As the concentration of additives does not affect cell viability, the supramolecular fibers established in this study are a promising candidate for various biomedical applications, such as standardized three-dimensional culture matrices for somatic cells and the regulation of stem cell differentiation.

Freestanding non-covalent thin films of the propeller-shaped polycyclic aromatic hydrocarbon decacyclene

Molecularly thin, nanoporous thin films are of paramount importance in material sciences. Their use in a wide range of applications requires control over their chemical functionalities, which is difficult to achieve using current production methods. Here, the small polycyclic aromatic hydrocarbon decacyclene is used to form molecular thin films, without requiring covalent crosslinking of any kind. The 2.5 nm thin films are mechanically stable, able to be free-standing over micrometer distances, held together solely by supramolecular interactions. Using a combination of computational chemistry and microscopic imaging techniques, thin films are studied on both a molecular and microscopic scale. Their mechanical strength is quantified using AFM nanoindentation, showing their capability of withstanding a point load of 26 ± 9 nN, when freely spanning over a 1 μm aperture, with a corresponding Young’s modulus of 6 ± 4 GPa. Our thin films constitute free-standing, non-covalent thin films based on a small PAH.

Vibrational Modes and Terahertz Phenomena of the Large-Cage Zeolitic Imidazolate Framework-71.

The zeolitic imidazole framework ZIF-71 has the potential to outperform other well-studied metal–organic frameworks due to its intrinsic hydrophobicity and relatively large pore size. However, a detailed description of its complex physical phenomena and structural dynamics has been lacking thus far. Herein, we report the complete assignment of the vibrational modes of ZIF-71 using high-resolution inelastic neutron scattering measurements and synchrotron radiation infrared spectroscopy, corroborated by density functional theory (DFT) calculations. With its 816 atoms per unit cell, ZIF-71 is the largest system yet for which frequency calculations have been accomplished employing the CRYSTAL17 DFT code. We discover low-energy terahertz dynamics such as gate-opening and shearing modes that are central to the functions and stability of the ZIF-71 framework structure. Nanoscale analytical methods based on atomic force microscopy (near-field infrared spectroscopy and AFM nanoindentation) further unravel the local chemical and mechanical properties of ZIF-71 single crystals.

Fourier Transform Infrared Microspectroscopy Combined with Principal Component Analysis and Artificial Neural Networks for the Study of the Effect of β-Hydroxy-β-Methylbutyrate (HMB) Supplementation on Articular Cartilage.

The potential of Fourier Transform infrared microspectroscopy (FTIR microspectroscopy) and multivariate analyses were applied for the classification of the frequency ranges responsible for the distribution changes of the main components of articular cartilage (AC) that occur during dietary β-hydroxy-β-methyl butyrate (HMB) supplementation. The FTIR imaging analysis of histological AC sections originating from 35-day old male piglets showed the change in the collagen and proteoglycan contents of the HMB-supplemented group compared to the control. The relative amount of collagen content in the superficial zone increased by more than 23% and in the middle zone by about 17%, while no changes in the deep zone were observed compared to the control group. Considering proteoglycans content, a significant increase was registered in the middle and deep zones, respectively; 62% and 52% compared to the control. AFM nanoindentation measurements collected from animals administered with HMB displayed an increase in AC tissue stiffness by detecting a higher value of Young’s modulus in all investigated AC zones. We demonstrated that principal component analysis and artificial neural networks could be trained with spectral information to distinguish AC histological sections and the group under study accurately. This work may support the use and effectiveness of FTIR imaging combined with multivariate analyses as a quantitative alternative to traditional collagenous tissue-related histology.

Load-dependent energy dissipation induced by the tip-membrane friction on suspended 2D materials.

The tip-membrane interface plays a critical role in characterizing the mechanical properties of ultrathin 2D materials by commonly employed nanoindentation based on atomic force microscopy (AFM). However, the reliability of the assumption that the tip-membrane interface remains pinned during nanoindentation remains unclear, which may introduce unignorable uncertainty in evaluating their true mechanical properties. In this work, it is reported that load-dependent frictional behavior would occur on the tip-membrane interface during nanoindentation tests on monolayer and multilayer suspended WS2 and graphene, and the curve hysteresis could be well explained by the stick-slip behavior. Further analyses and finite element simulations demonstrated that the frictional energy dissipation should be mainly attributed to the frictional behavior along the direction parallel to the cantilever beam. Meanwhile, the in-plane membrane stiffness was mainly responsible for the different frictional behavior on monolayer and multilayer 2D materials. Based on these analyses, some suggestions were proposed to help reduce the uncertainty when extracting the mechanical properties of 2D materials. These findings not only facilitate the deep understanding of the origin of the curve hysteresis during nanoindentation, but also help to evaluate the mechanical properties of 2D materials in a more reliable way.

Mechanical rigidity of a shape-memory metal-organic framework increases by crystal downsizing.

Soft porous nanocrystals with a pronounced shape-memory effect exhibit two- to three-fold increase in elastic modulus compared to the microcrystalline counterpart as determined by atomic force microscopy nanoindentation. The increase in rigidity is consistent with the known shape-memory effect displayed by the framework solid at the nanoscale. Crystal downsizing can offer new avenues for tailoring the mechanical properties of metal-organic frameworks.

Lignin-based nanogels for the release of payloads in alkaline conditions

Bio-based nanoparticles are promising materials in agriculture because they enable a controlled release of agrochemicals to plants. The release can be more specific by engineering nanoparticles with stimuli-responsive properties. Herein, we reported lignin-based nanogels which can release payloads in alkaline conditions. Lignin methacrylate was emulsified in miniemulsion droplets and graft copolymerized with acrylic acid to form core-shell nanogels. The successful incorporation of acrylic acid and lignin units in the nanogels was confirmed by FT-IR, H-NMR and XPS spectroscopies. TEM and dynamic light scattering analysis demonstrated that the nanogels were spherical and uniformly dispersed in the aqueous phase. AFM nanoindentation and thermal analysis indicated that the nanogels were mechanically and thermally durable. Lignin-based nanogels exhibited a pH-responsive swelling at high amounts of acrylic acid. Various hydrophobic payloads were encapsulated in high yield and could be preferentially released in alkaline conditions. Lignin-based nanogels are therefore the potential nanocarrier for the controlled release of agrochemicals such as pesticides or fungicides.

Nanomechanical properties of Monilethrix affected hair are independent of phenotype

Utilising the AFM nanoindentation technique for the study of hair cross- and longitudinal sections, the mechanical anisotropy of human hair fibres affected by a rare congenital condition, Monilethrix, has been investigated for the first time. Supported by X-ray microdiffraction data, and applying a model based on an ideal composite material consisting of rods (KIFs) and matrix (KAPs) to Monilethrix affected fibres, it has been shown that the results could be grouped into clearly different classes, namely: almost isotropic behaviour for Monilethrix affected hairs and anisotropic behaviour for Control hair. Moreover, AFM nanoindentation of hair cross sections has demonstrated, also for the first time that hairs affected by Monilethrix have a continuous, and not periodic, weakness within the cortex. This has been attributed to disruptions in the KIF-KIF, KIF-intermacrofibrillar matrix or KIF-desmosome complexes within the hair shaft, as suggested by X-ray microdiffraction examination. Hairs from a patient exhibiting no obvious phenotype exhibited similar mechanical weakness despite the otherwise normal visual appearance of the fibre. This further supports a hypothesis that the beaded appearance of Monilethrix hair is a secondary factor, unrelated to the inherent structural weakness.

(Invited) Instability and Inhomogeneity in the Lithiation of Methylated Amorphous Silicon Layers

Silicon is one of the best candidates as an anode material for Li-ion batteries, owing to its high capacity. However, the large variation in volume induced by lithiation leads to a degradation of the material incompatible with its practical use. We have shown that amorphous methylated silicon a-Si1− x (CH3) x :H has a better cyclability while keeping a comparable capacity [1]. This was attributed to a lower stiffness of the methylated material as compared to pure silicon. AFM nanoindentation experiments confirm this property.The first lithiation/delithiation cycles are of prime importance for the evolution of the material during subsequent cycling. Operando monitoring by attenuated-total-reflection Fourier-transform infrared spectroscopy and optical microscopy provide a thorough understanding of the mechanisms involved during these cycles. The lithiation of a-Si1− x (CH3) x :H, with various methyl contents (x = 0 - 0.12), was investigated using operando ATR-FTIR [2]. The first lithiation proceeds via a two-phase mechanism [3], whatever the methyl content is. The lithium concentration z of the invading Li-rich phase depends on x, first decreasing for x < 0.05 and increasing above (Fig.1). This behavior was tentatively explained by two distinct effects: the softening of the material due to a methyl-induced lowering of its reticulation degree and its cohesion, and the presence of nanovoids at higher methyl content.Operando observations by optical microscopy reveal that, unlike pure silicon, the methylated silicon undergoes a first spatially inhomogeneous lithiation [4]: lithiated zones appear on the surface as colored spots (Fig.2), which grow circularly and progressively invade the surface. These zones have a thickness greater than that of the initial layer, which can be estimated from their color change. The morphology of the lithiation spots and their evolution are accurately determined by ex situ AFM. The appearance of these lithiation spots appears to be related to an electrostatic instability associated with the resistive character of the a-Si1− x (CH3) x :H layer. This process is reminiscent of that responsible for the formation of macropores on p-Si, but at a much larger length scale [5]. Experimental observations unravel the existence of defects determining the locations at which lithiation spots are formed. A simple model has been worked out for the lateral growth of the lithiation spots, which is in semi-quantitative agreement with the combined electrochemical and microscopy measurements. Complementary Raman spectroscopy experiments give some insight into the structural and electrochemical changes during the first lithiation/delithiation cycle.[1] L. Touahir, A. Cheriet, D.A. Dalla Corte, J.-N. Chazalviel, C. Henry de Villeneuve, F. Ozanam, I. Solomon, A. Keffous, N. Gabouze, M. Rosso, J. Power Sources. 240 (2013) 551–557.[2] B.M. Koo, D.A. Dalla Corte, J.-N. Chazalviel, F. Maroun, M. Rosso, F. Ozanam, Adv. Energy Mater. 8 (2018) 1702568.[3] J.W. Wang, Y. He, F. Fan, X.H. Liu, S. Xia, Y. Liu, C.T. Harris, H. Li, J.Y. Huang, S.X. Mao, T. Zhu, Nano Lett. 13 (2013) 709–715; M.T. McDowell, S.-W. Lee, J.T. Harris, B.A. Korgel, C. Wang, W.D. Nix, Y. Cui, Nano Lett 13 (2013) 758–764.[4] Y. Feng, T.-D.-T. Ngo, M. Panagopoulou, A. Cheriet, B.M. Koo, C. Henry-de-Villeneuve, M. Rosso, F. Ozanam, Electrochim. Acta 302 (2019) 249-258.[5] J.-N. Chazalviel, F. Ozanam, N. Gabouze, S. Fellah, R. B. Wehrspohn, J. Electrochem. Soc. 149 (2002) C511-C520. Figure 1

A New Approach for the AFM-Based Mechanical Characterization of Biological Samples.

The AFM nanoindentation technique is a powerful tool for the mechanical characterization of biological samples at the nanoscale. The data analysis of the experimentally obtained results is usually performed using the Hertzian contact mechanics. However, the aforementioned theory can be applied only in cases that the sample is homogeneous and isotropic and presents a linear elastic response. However, biological samples often present depth-dependent mechanical properties, and the Hertzian analysis cannot be used. Thus, in this paper, a different approach is presented, based on a new physical quantity used for the determination of the mechanical properties at the nanoscale. The aforementioned physical quantity is the work done by the indenter per unit volume. The advantages of the presented analysis are significant since the abovementioned magnitude can be used to examine if a sample can be approximated to an elastic half-space. If this approximation is valid, then the new proposed method enables the accurate calculation of Young's modulus. Additionally, it can be used to explore the mechanical properties of samples that are characterized by a depth-dependent mechanical behavior. In conclusion, the proposed analysis presents an accurate yet simple technique for the determination of the mechanical properties of biological samples at the nanoscale that can be also used beyond the Hertzian limit.

Differential flexibility leading to crucial microelastic properties of asymmetric lipid vesicles for cellular transfection: A combined spectroscopic and atomic force microscopy studies

The role of microscopic elasticity of nano-carriers in cellular uptake is an important aspect in biomedical research. Herein we have used AFM nano-indentation force spectroscopy and Förster resonance energy transfer (FRET) measurements to probe microelastic properties of three novel cationic liposomes based on di-alkyl dihydroxy ethyl ammonium chloride based lipids having asymmetry in their hydrophobic chains (Lip1818, Lip1814 and Lip1810). AFM data reveals that symmetry in hydrophobic chains of a cationic lipid (Lip1818) imparts higher rigidity to the resulting liposomes than those based on asymmetric lipids (Lip1814 and Lip1810). The stiffness of the cationic liposomes is found to decrease with increasing asymmetry in the hydrophobic lipid chains in the order of Lip1818 > Lip1814 > lip1810. FRET measurements between Coumarin 500 (Donor) and Merocyanine 540 (Acceptor) have revealed that full width at half-maxima (hw) of the probability distribution (P(r)) of donor-acceptor distance (r), increases in an order Lip1818 < Lip1814 < Lip1810 with increasing asymmetry of the hydrophobic lipid chains. This increase in width (hw) of the donor-acceptor distance distributions is reflective of increasing flexibility of the liposomes with increasing asymmetry of their constituent lipids. Thus, the results from AFM and FRET studies are complementary to each other and indicates that an increase in asymmetry of the hydrophobic lipid chains increases elasticity and or flexibility of the corresponding liposomes. Cell biology experiments confirm that liposomal flexibility or rigidity directly influences their cellular transfection efficiency, where Lip1814 is found to be superior than the other two liposomes manifesting that a critical balance between flexibility and rigidity of the cationic liposomes is key to efficient cellular uptake. Taken together, our studies reveal how asymmetry in the molecular architecture of the hydrophobic lipid chains influences the microelastic properties of the liposomes, and hence, their cellular uptake efficiency.

One-step process for the preparation of fast-response soft actuators based on electrospun hybrid hydrogel nanofibers obtained by reactive electrospinning with in situ synthesis of conjugated polymers

Soft actuators with a fast response to low electrical potentials are critical for wearables and robotics. In this work, hybrid hydrogel nanofiber mats were prepared and tested as soft actuators. Hybrid hydrogel nanofibers were produced by combining electrospinning with in situ photopolymerization of the hydrogel and polymerization of aniline during the formation of the nanofibers. The electrical conductivity of the obtained nanofibers measured by the 4-probe method reached values close to 0.01 S cm−1. Images obtained by atomic force microscopy in conductive mode (I-AFM) were also successfully acquired and proved the conductive and nanometric nature of the hybrid hydrogel nanofibers. Infrared spectra (FTIR) showed that photopolymerization of the hydrogel precursors and aniline polymerization during nanofiber formation were effectively achieved. The elastic modulus of the nanofibers was estimated from AFM nanoindentations, and values close to 10 MPa for the swollen samples were obtained. The swelling ability of the hybrid hydrogel nanofibers was evaluated by comparing AFM images of the nanofibers in dried and swollen conditions to reveal that the swelling degree is reduced by the presence of polyaniline in the nanofibers. The obtained hybrid hydrogel nanofiber mats were tested as actuators in an electrochemical cell. The application of low voltage (1 V) in a 1 MΩ cm water led to fast (2.5 mm s−1) and reversible displacement of the samples. During actuation, thin nanofiber mats could also withstand forces on the order of 80 μN that were measured by coupling the electrochemical cell to a load cell.

Nucleation and Growth of Lithiation Patterns on Methylated Amorphous Silicon Layers Monitored By Operando Optical Microscopy

Silicon is one of the best candidates as an anode material for Li-ion batteries, owing to its high capacity. However, the large variation in volume induced by lithiation leads to a degradation of the material incompatible with its use in practice. We have shown that amorphous methylated silicon a-Si 1-x (CH3) x :H has a better cyclability while keeping a comparable capacity [1]. This was attributed to a lower stiffness of the methylated material as compared to pure silicon. AFM nanoindentation experiments confirm this property.The first lithiation/delithiation cycles are of prime importance for the evolution of the material during subsequent cycling. Operando monitoring by attenuated-total-reflection Fourier-transform infrared (ATR-FTIR) spectroscopy and optical microscopy provide a thorough understanding of the mechanisms involved during these cycles. The lithiation of a-Si 1-x (CH3) x :H, with various methyl contents (x = 0 - 0.12), was investigated using operando ATR-FTIR [2]. As previously reported for pure crystalline and amorphous silicon [3], the first lithiation proceeds via a two-phase mechanism whatever the methyl content is. The lithium concentration z of the invading Li-rich phase depends on x, first decreasing for x < 0.05 and increasing above (Fig.1). This behavior was tentatively explained by two distinct effects: the softening of the material due to a methyl-induced lowering of its reticulation degree and its cohesion, and the presence of nanovoids at higher methyl content.Operando observations by optical microscopy were performed on thin layers during the first lithiation. Unlike pure silicon, the methylated silicon undergoes a first spatially inhomogeneous lithiation [4]: lithiated zones appear on the surface as colored spots (Fig.2), which grow circularly and progressively invade the surface. These spots have a thickness greater than that of the initial layer, which can be estimated from their color change. The morphology of the lithiation spots and their evolution are accurately determined by ex situ AFM. The resistive character of the layer is found to be responsible for this inhomogeneous behavior. A simple model has been worked out and is in semi-quantitative agreement with the combined electrochemical and microscopy measurements.Using Raman spectroscopy, we investigated the properties of fully lithiated, semi-lithiated and non-lithiated layers, in and out of the lithiation spots. Our results give some insight into the structural and electrochemical changes during the first cycle. They also show the low stability of the material under illumination and electrochemical conditions.

Micropatterned Biphasic Nanocomposite Platform for Maintaining Chondrocyte Morphology.

One major limitation hindering the translation of in vitro osteoarthritis research into clinical disease-modifying therapies is that chondrocytes rapidly spread and dedifferentiate under standard monolayer conditions. Current strategies to maintain rounded morphologies of chondrocytes in culture either unnaturally restrict adhesion and place chondrocytes in an excessively stiff mechanical environment or are impractical for use in many applications. To address the limitations of current techniques, we have developed a unique composite thin-film cell culture platform, the CellWell, to model articular cartilage that utilizes micropatterned hemispheroidal wells, precisely sized to fit individual cells (12-18 μm diameters), to promote physiologically spheroidal chondrocyte morphologies while maintaining compatibility with standard cell culture and analytical techniques. CellWells were constructed of 15-μm-thick 5% agarose films embedded with electrospun poly(vinyl alcohol) (PVA) nanofibers. Transmission electron microscope (TEM) images of PVA nanofibers revealed a mean diameter of 60.9 ± 24 nm, closely matching the observed 53.8 ± 29 nm mean diameter of human ankle collagen II fibers. Using AFM nanoindentation, CellWells were found to have compressive moduli of 158 ± 0.60 kPa at 15 μm/s indentation, closely matching published stiffness values of the native pericellular matrix. Primary human articular chondrocytes taken from ankle cartilage were seeded in CellWells and assessed at 24 h. Chondrocytes maintained their rounded morphology in CellWells (mean aspect ratio of 0.87 ± 0.1 vs three-dimensional (3D) control [0.86 ± 0.1]) more effectively than those seeded under standard conditions (0.65 ± 0.3), with average viability of >85%. The CellWell's design, with open, hemispheroidal wells in a thin film substrate of physiological stiffness, combines the practical advantages of two-dimensional (2D) culture systems with the physiological advantages of 3D systems. Through its ease of use and ability to maintain the physiological morphology of chondrocytes, we expect that the CellWell will enhance the clinical translatability of future studies conducted using this culture platform.