Nanoindentation Protocols Research Articles

Dynamic Nanoindentation and Short-Range Order in Equiatomic NiCoCr Medium-Entropy Alloy Lead to Novel Density Wave Ordering.

Chemical short-range order is believed to be a key contributor to the exceptional properties of multicomponent alloys. However, its direct validation and confirmation has been highly elusive in most compounds. Recent studies for equiatomic NiCoCr alloys have shown that thermal treatments (i.e., annealing and aging) may facilitate and manipulate such ordering. In this work, by using molecular simulations, we show that nanomechanical probes, such as nanoindentation, may be utilized toward further manipulation of chemical short-range order, providing explicit validation pathways. By using well established interatomic potentials, we perform hybrid molecular-dynamics-MonteCarlo at room temperature to demonstrate that particular dwell nanoindentation protocols can lead, through thermal MonteCarlo equilibration, to local reorganization under the indenter tip, toward a density-wave stripe pattern. We characterize the novel density-wave structures, which are highly anisotropic and dependent on local, nanoindentation-induced stress concentrations, and we show how they deeply originate from intrinsic features of interelemental interactions. Furthermore, we show that these novel patterns consistently scale with the incipient plastic zone, under the indenter tip, justifying their observation at experimentally feasible nanoindentation depths.

Micromechanics of non-embedded spruce wood: Novel polishing and indentation protocol

Since the start of nanoindentation in wood, cell walls have been supported by epoxy resin during the microtoming of surfaces. However, wood embedding in epoxy resin was initially developed to harvest thin sections destined for transmission electron microscopy. In nanoindentation, embedding can affect the identified mechanical properties and might be difficult for modified wood samples. For this reason, we propose a protocol that does not involve resin within the wood specimen, uses a more robust miller to achieve surfaces with root means squared roughness of around 35 nm, and can exclude invalid edge indents through a novel evaluation procedure based on fitting 2D histograms with a linear combination of bi-variate lognormal probability density functions. We examined twelve samples of Norway spruce (Picea abies) late wood and their counterparts soaked with a biological preservation pretreatment. A nanoindentation protocol on every sample led to a mean reduced modulus and mean hardness of around 12 % and 26 %, respectively, lower than those found in the literature for embedded Norway spruce. These results agree well with previous results found for Loblolly pine (Pinus taeda L.) when comparing embedded and non-embedded specimens.

Deformation behaviors of a model metallic glass under 3-D nanoindentation studied in molecular dynamics simulation

In recent years, molecular dynamics (MD) simulation has been used to study the deformation behaviors of glass under nanoindentation, mainly using ideal geometries like a spherical indenter or a 2.5-D sample geometry to simplify post-analysis and save computational costs. To generate stress/strain fields that can be directly compared with experiments, we developed a 3-D nanoindentation protocol in this work to study the deformation behaviors of a model metallic glass under sharp contact loading in MD. Our studies show that the indenter sharpness controls the shear band formation, and the interaction between shear bands dictates the crack initiation in the model metallic glass. Shear bands and residual stress fields in the model metallic glass from our simulated nanoindentation tests are consistent with observations in soda-lime silicate (SLS) glass from the instrumented indentation in experiments, as both of them favor shear deformation under sharp contact loading.

Nano-scale fracture toughness of fly ash incorporated hydrating cementitious composites using experimental nanoindentation technique

Instrumented nanoindentation technique has proven its strength by providing incredible insight into the material microstructure of homogeneous materials. Its success has led to its implementation in heterogenous materials such as cementitious composites. The implementation of instrumented nanoindentation on cement paste composites has proven to be very fruitful as it has enabled the evaluation of fundamental properties including Young’s modulus, hardness, creep at multiscales. However, there are more complex properties like fracture toughness which are crucial to characterize the material at macro scales. This characterisation would be benefitted by studies at micro-, nano- scales of the complex properties; but they are seldom reported. Cement-based materials have extremely complex, multiscale structures that are initiated from the chemical reactions of the hydration process and then continue to evolve with time. Also, preparing the surface of hydrating cementitious composites requires special consideration as it can greatly affect the indentation results. To explore the microstructural composition and its fracture toughness, an experimental grid nanoindentation protocol has been put forward. This experimental protocol has been implemented on three cement pastes, i.e., plain Ordinary Portland Cement (OPC), OPC + 20% fly ash, and OPC + 40% fly ash during their early stages as well as near complete stages of hydration. Since measuring the micro cracks around the indent is virtually near impossible, the load vs. displacement curves recorded by the indentations have been utilized in an energy-based method to evaluate the energy release rate and fracture toughness. The evaluated values of fracture toughness property of the individual microstructural phases will help in shedding a light on the development and spread of fracture toughness in cement-based materials. In this study the different methods for fracture toughness evaluation at the micro scales using nanoindentation technique are reported on and the method suitable for cementitious materials is implemented to evaluate fracture toughness of the microstructural phases.

Evolution of the viscoelastic properties of painting stratigraphies: a moisture weathering and nanoindentation approach

In this investigation on the mechanical behaviour of paint films, we use a new ferrule-top nanoindentation protocol developed for cultural heritage studies to examine the impact of repeated relative humidity variations on the viscoelastic behaviour of paint films and their mechanical properties in different paint stratigraphies through the changes in their storage and loss moduli. We show that the moisture weathering impact on the micromechanics varies for each of these pigment-oil systems. Data from the nanoindentation protocol provide new insights into the evolution of the viscoelastic properties dsue to the impact of moisture weathering on paint films.

The use of ground glass in red glazes: structural 3D imaging and mechanical behaviour using optical coherence tomography and nanoindentation

In this study we investigate the impact of the addition of colourless glass particles to red glazes, as seen in many 15th-17th-century easel paintings. With the use of reconstructions, we examined the influence of the paint preparation process on the morphological and mechanical properties of the paint film. Three sets of reconstructions were made, a control without ground glass, reconstructions with coarse or fine ground glass mixed in, and reconstructions where fine ground glass was ground jointly with the pigment oil mixture. The latter gave the desired consistency and colour based on visual inspection. The dried reconstructions were non-invasively imaged using optical coherence tomography (OCT). A data-analysis pipeline was developed for both the segmentation of the OCT images and the measurement of the size and spatial distributions of the glass particles within the glaze layer. Moreover, we used a nanoindentation protocol to measure the viscoelastic properties of the dried red glaze film. The OCT results show an expected decrease in median particle size with longer grinding-time, for which the additional grinding with pigment/oil resulted in a more narrow size distribution and a homogenous spatial distribution of the glass particles. The nanoindentation results indicate that the addition of glass particles increases the elastic and viscous moduli of the red glaze layers. The homogeneous size distribution, obtained by grinding the oil, pigment, and glass together, induces higher elastic and viscous moduli. Our imaging and analyses approach, combining OCT and nanoindentation, provides a non-invasive and quantitative investigation of glass particles in (semi-) transparent paint layers, and their effect on the mechanical properties of the glaze. The results of this study contribute to a better understanding of the artists’ addition of ground glass in paint layers.

A novel nanoindentation protocol to characterize surface free energy of superhydrophobic nanopatterned materials

Surface Free Energy (SFE) has become a relevant design parameter to produce materials and devices with controlled wettability. The non-destructive measurement of SFE in nanopatterned super-hydrophobic hard surfaces is a challenge in both research and industry since in most cases time-consuming contact angle measurements are not feasible. In this work, we present a novel nanoindentation based method for the measurement of pull-off adhesive forces by carefully controlling environmental and instrumentation issues. The method is found to measure SFE over five orders of magnitude, covering hydrophilic to super-hydrophobic surfaces, and has been validated with contact angle measurements. Its limitations and shortcomings are critically discussed, with a specific focus on the experimental issues that could affect the reliability and reproducibility of the results. Finally, the potential applications of the newly developed methodology include fast non-destructive mapping of SFE over heterogeneous surfaces with spatially controlled wettability.Graphic abstract

Cell spheroid fusion: beyond liquid drops model

Biological self-assembly is crucial in the processes of development, tissue regeneration, and maturation of bioprinted tissue-engineered constructions. The cell aggregates—spheroids—have become widely used model objects in the study of this phenomenon. Existing approaches describe the fusion of cell aggregates by analogy with the coalescence of liquid droplets and ignore the complex structural properties of spheroids. Here, we analyzed the fusion process in connection with structure and mechanical properties of the spheroids from human somatic cells of different phenotypes: mesenchymal stem cells from the limbal eye stroma and epithelial cells from retinal pigment epithelium. A nanoindentation protocol was applied for the mechanical measurements. We found a discrepancy with the liquid drop fusion model: the fusion was faster for spheroids from epithelial cells with lower apparent surface tension than for mesenchymal spheroids with higher surface tension. This discrepancy might be caused by biophysical processes such as extracellular matrix remodeling in the case of mesenchymal spheroids and different modes of cell migration. The obtained results will contribute to the development of more realistic models for spheroid fusion that would further provide a helpful tool for constructing cell aggregates with required properties both for fundamental studies and tissue reparation.

Process-Structure-Property Modeling for Severe Plastic Deformation Processes Using Orientation Imaging Microscopy and Data-Driven Techniques

Machining is a severe plastic deformation process, wherein the workpiece material is subjected to high deformation rates and temperatures. During metal machining, the dynamic recrystallization mechanism causes grain refinement into the sub-micron range. In this study, we investigate the microstructure evolution of oxygen-free high conductivity copper (OFHC Cu) subject to a machining process where the cutting speed and rake angle are controlled to manipulate the process strain, strain rate, and temperatures. Microstructures of the deformed chips are quantified using orientation imaging microscopy and novel statistical descriptors that capture the morphology and local lattice misorientations generated during the several mechanistic stages of the dynamic recrystallization process. Mechanical properties of the resulting chips are quantified using spherical nanoindentation protocols. A multiple output Gaussian process regression model is used to simultaneously model the structure-property evolution, which differs from more common approaches that establish such relationships sequentially. This modeling strategy is particularly attractive since it can flexibly provide both structure and property uncertainty estimates. In addition, the statistical modeling framework allows for the inclusion of multi-fidelity data. The statistical metrics utilized serve as efficient microstructure descriptors, which retain the physics of the observed structures without having to introduce ad hoc microstructure feature definitions.

High Temperature Flow Behavior of Ultra-Strong Nanoporous Au assessed by Spherical Nanoindentation.

Nanoporous metals have attracted attention in various research fields in the past years since their unique microstructures make them favorable for catalytic, sensory or microelectronic applications. Moreover, the refinement of the ligaments down to the nanoscale leads to an exceptionally high strength. To guarantee a smooth implementation of nanoporous metals into modern devices their thermo-mechanical behavior must be properly understood. Within this study the mechanical flow properties of nanoporous Au were investigated at elevated temperatures up to 300 °C. In contrast to the conventional synthesis by dealloying of AuAg precursors, the present foam was fabricated via severe plastic deformation of an AuFe nanocomposite and subsequent selective etching of iron, resulting in Au ligaments consisting of nanocrystalline grains, while remaining Fe impurities excessively stabilize the microstructure. A recently developed spherical nanoindentation protocol was used to extract the stress-strain curves of nanoporous Au. A tremendous increase of yield strength due to ligament and grain refinement was observed, which is largely maintained at high temperatures. Reviewing literature will evidence that the combined nanocrystalline and nanoporous structure leads to remarkable mechanical properties. Furthermore, comparison to a previous Berkovich nanoindentation study outlines the conformity of different indentation techniques.

Type 2 Diabetic Coronary Vascular Smooth Muscle Cells Exhibit Decreased Stiffness and Decreased Adhesion

We previously reported that type 2 diabetic (T2D) coronary resistance microvessels (CRMs) undergo early inward hypertrophic remodeling with reduced vessel stiffness that is associated with a proliferative VSMC phenotype. We also previously revealed that there is no difference in stiffness of decellularized CRMs from diabetic (db/db) and normal (Db/db) mice, suggesting that the decreased stiffness of the intact diabetic CRM is not due to alterations in the extracellular matrix (ECM) of those microvessels. Given that diabetic CRMs are less stiff and this decreased stiffness does not appear to be due differences in the mechanical strength of the surrounding ECM, the aim of the current study was to determine whether a reduction in cellular mechanical stiffness between Db/db and db/db CRM vascular smooth muscle cells (VSMCs) may be responsible. VSMCs were isolated from 16 week old Db/db and db/db mice and passage 1 cells were used for all experiments (n=6–7 for all groups). Using an atomic force microscope with probes coated with fibronectin (FN), a nano‐indentation protocol was used to measure cellular stiffness and adhesion to FN. Elastic modulus, a measurement of cellular stiffness, was reduced in diabetic coronary VSMCs compared to normal (Db/db: 6.74 ± 0.40 kPa vs. db/db: 4.96 ± 0.35 kPa, p = 0.004). By comparison, there were no significant differences in elastic modulus between normal and diabetic aortic VMSCs (Db/db: 5.39 ± 0.79 kPa vs. db/db: 5.18 ± 0.87 kPa, p = 0.861). Additionally, diabetic coronary VSMCs had reduced adhesive force to fibronectin compared to normal coronary VSMCs (Db/db: 54.78 ± 5.49 vs. db/db: 39.8 ± 3.43 pN, p = 0.03). In a separate experiment, qPCR was performed on mRNA from normal and diabetic CRM VSMCs. Alpha‐smooth muscle actin trended lower in db/db CRM VSMCs (p=0.07), indicating a de‐differentiated phenotype. Cell cycle regulator, p53, had significantly reduced mRNA expression in the diabetic coronary VSMCs over control VSMCs (Db/db 1.00±0.06 vs. db/db 0.63±0.08, p<0.01). These data show that reductions in cellular stiffness and adhesion to the ECM of diabetic CRM VSMCs may underlie ex vivo whole microvessel mechanical properties to account for reduced CRM stiffness. The changes in diabetic coronary VSMC stiffness and adhesion may be the result of VMSC phenotypic switching to a more de‐differentiated/proliferative phenotype.Support or Funding InformationSupport: NIH R00 HL116769 and S10 OD023438 and Nationwide Children's Hospital to AJTThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Essential refinements of spherical nanoindentation protocols for the reliable determination of mechanical flow curves

Understanding and linking mechanical properties obtained by spherical indentation experiments to uniaxial data is extremely challenging. Since the first attempts in the early 20th century numerous advances gradually allowed to expand the output of indentation tests. Still, the extraction of flow curves from spherical nanoindentation has not yet been fully established, as tip shape problems and size effects impede a straight-forward implementation. Within this study, we show new calibration procedures originating from fundamental geometrical considerations to account for tip shape imperfections. This sets the base for strain-rate controlled tests, which in turn enables us to measure rate-dependent material properties either with constant strain-rate or by strain-rate jump tests. Finally, experimental evaluation of the constraint factor in consideration of the mechanical properties and induced strain enables the extraction of flow curves. Testing materials with refined microstructures ensures the absence of possible size-effects. This study contributes to a significant improvement of current experimental protocols and allows to move flow curve measurements from single spherical imprints one step closer to its implementation as a standard characterization technique for modern materials.

Quantifying the mechanical effects of He, W and He + W ion irradiation on tungsten with spherical nanoindentation

Recent advances in spherical nanoindentation protocols have proven very useful for capturing the grain-scale mechanical response of different metals. This is achieved by converting the load–displacement response into an effective indentation stress–strain response which reveals latent information such as the elastic–plastic transition or indentation yield strength and work-hardening behavior and subsequently correlating the response with the material structure (e.g., crystal orientation) at the indentation site. Using these protocols, we systematically study and quantify the microscale mechanical effects of He, W, and He + W ion irradiation on commercially pure, polycrystalline tungsten. The indentation stress–strain response is correlated with the crystal orientation from electron backscatter diffraction, the defect structure from transmission electron microscopy micrographs, and the stopping range of ions in matter calculations of displacement damage and He concentration. He-implanted grains show a much higher indentation yield strength and saturation stress compared to W-ion-irradiated grains for the same displacement damage. There is also good agreement between the dispersed barrier hardening model with a barrier strength of 0.5–0.8 and void models (Bacon–Kochs–Scattergood and Osetsky–Bacon models) with the experimentally observed changes in indentation strength due to the presence of He bubbles. This finding indicates that a high density (~ 9 × 1023 m−3) and concentration (~ 1.5 at.%) of small (~ 1 nm diameter) He bubbles can be moderate to strong barriers to dislocation slip in tungsten.

New methods for the evaluation of bone quality. Nanoindentation protocol for measurement of bone mechanical properties of material level.

Bone is an inhomogeneous, anisotropic natural biomaterial with complex, multiscale structural variations. Thus, experiments on the bulk scale using a universal testing machine are not applicable for localized precision mechanical testing of bone. Nanoscale mechanical testing technologies such as nanoindentation enables to assess the intrinsic toughening mechanism of bone, which is a function of the highly-organized matrix proteins within the mineralized nanostructure. Understanding the basic nanomechanical properties of calcified tissues will help us to appreciate general concepts associated with the excellent design of advanced engineering materials and engineered tissues.

High throughput exploration of process-property linkages in Al-6061 using instrumented spherical microindentation and microstructurally graded samples.

Recent spherical nanoindentation protocols have proven robust at capturing the local elastic-plastic response of polycrystalline metal samples at length scales much smaller than the grain size. In this work, we extend these protocols to length scales that include multiple grains to recover microindentation stress-strain curves. These new protocols are first established in this paper and then demonstrated for Al-6061 by comparing the measured indentation stress-strain curves with the corresponding measurements from uniaxial tension tests. More specifically, the scaling factors between the uniaxial yield strength and the indentation yield strength was determined to be about 1.9, which is significantly lower than the value of 2.8 used commonly in literature. The reasons for this difference are discussed. Second, the benefits of these new protocols in facilitating high throughput exploration of process-property relationships are demonstrated through a simple case study.

A New Nanoindentation Protocol for Identifying the Elasticity of Undamaged Extracellular Bone Tissue

ABSTRACTWhile the quest for understanding and even mimicking biological tissue has propelled, over the last decades, more and more experimental activities at the micro and nanoscales, the appropriate evaluation and interpretation of respective test results has remained a formidable challenge. As a contribution to tackling this challenge, we here describe a new method for identifying, from nanoindentation, the elasticity of the undamaged extracellular bone matrix. The underlying premise is that the tested bovine bone sample is either initially damaged (i.e. exhibits micro-cracks a priori) or that such micro-cracks are actually induced by the nanoindentation process itself, or both. Then, (very many) indentations may relate to either an intact material phase (which is located sufficiently far away from micro-cracks), or to differently strongly damaged material phases. Corresponding elastic phase properties are identified from the statistical evaluation of the measured indentation moduli, through representation of their histogram as a weighted sum of Gaussian distribution functions. The resulting undamaged elastic modulus of bovine femoral extracellular bone matrix amounts to 31 GPa, a value agreeing strikingly well both with direct quasi-static modulus tests performed on SEM-FIB-produced micro-pillars (Luczynski et al., 2015), and with the predictions of a widely validated micromechanics model (Morin and Hellmich, 2014). Further confidence is gained through observing typical indentation imprints under Scanning Electron Microscopy (SEM), which actually confirms the existence of the two types of micro-cracks as described above.

Possibility for rapid generation of high-pressure phases in single-crystal silicon by fast nanoindentation

High-pressure phases of silicon such as Si-XII/Si-III exhibit attractive optical, electrical and chemical properties, but until now, it has been technologically impossible to produce a significant quantity of Si-XII or Si-III. In this study, to explore the possibility of generating high-pressure silicon phases efficiently, comparative nanoindentation experiments were conducted. Effects of the loading rate, unloading rate and maximum indentation load were investigated, and key factors affecting the high-pressure phase formation were identified. A new nanoindentation protocol is proposed that introduces an intermediate holding stage into the unloading process. The resulting end phases under the indent were detected by a laser micro-Raman spectrometer and compared with those formed in conventional nanoindentation. The results indicate that high-pressure phases Si-XII and Si-III were successfully formed during the intermediate holding stage even with a very high loading/unloading rate. This finding demonstrates the possibility of rapid production of high-pressure phases of silicon through fast mechanical loading and unloading.

Time-dependent mechanical properties of rat femoral cortical bone by nanoindentation: An age-related study

Abstract

Increased Adhesion of Glycated Proteins to Arteriolar Vascular Smooth Muscle Cells as Determined by Atomic Force Microscopy

Modification of proteins by non-enzymatic glycation occurs in diabetes mellitus, aging and pathological states including renal failure. Glycation reactions span early reversible reactions to irreversibly modified species termed advanced glycation endproducts (AGEs). Long-lived extracellular matrix (ECM) proteins are particularly susceptible to AGE modification. As ECM proteins are known to regulate vascular function through binding with integrins, AGE formation may modify normal interactions between matrix and vascular smooth muscle cells (VSMCs) ultimately impacting vasoregulation. The purpose of this study was to selectively investigate the adhesion between either non-glycated or glycated fibronectin (FN) with VSMCs using Atomic Force Microscopy (AFM). AFM probes were functionalized with FN or glycated FN and cyclically interacted with isolated VSMCs using a force mode nano-indentation protocol. Adhesion was quantified as the rupture force required to disrupt adhesion bonds the formed during contact of the AFM probe with the cell and also as the probability of an adhesive interaction occurring. AFM probes coated with FN alone required 78.6 ± 19.9 pN (N=5) to disrupt adhesive bonds whereas glycated FN required 49.9 ± 5.7 pN (N=5). Binding probability was, however, significantly increased for AFM probes coated with glycated FN vs FN alone, 72.9 ± 3.5 % vs 63.0 ± 1.6 %, (N=5) respectively. Selectivity for binding to alpha5 integrin was investigated by adding a solubilized function-blocking antibody to the cell bath. Anti-alpha5 antibody significantly reduced binding to FN (14.27%) but had no effect on glycated FN. These results establish that glycation of ECM proteins can significantly affect their adhesive interaction with VSM and integrins and this could in turn lead to altered cell-integrin based regulation of VSM function. (Supp by P01 HL095486 and RO1 HL92241)

Precision of nanoindentation protocols for measurement of viscoelasticity in cortical and trabecular bone

Nanoindentation has recently gained attention as a characterization technique for mechanical properties of biological tissues, such as bone, on the sub-micron level. However, optimal methods to characterize viscoelastic properties of bones are yet to be established. This study aimed to compare the time-dependent viscoelastic properties of bone tissue obtained with different nanoindentation methods. Bovine cortical and trabecular bone samples (n=8) from the distal femur and proximal tibia were dehydrated, embedded and polished. The material properties determined using nanoindentation were hardness and reduced modulus, as well as time-dependent parameters based on creep, loading-rate, dissipated energy and semi-dynamic testing under load control. Each loading protocol was repeated 160 times and the reproducibility was assessed based on the coefficient of variation (CV). Additionally, three well-characterized polymers were tested and CV values were calculated for reference.The employed methods were able to characterize time-dependent viscoelastic properties of bone. However, their reproducibility varied highly (CV 9–40%). The creep constant increased with increasing dwell time. The reproducibility was best with a 30s creep period (CV 18%). The dissipated energy was stable after three repeated load cycles, and the reproducibility improved with each cycle (CV 23%). The viscoelastic properties determined with semi-dynamic test increased with increase in frequency. These measurements were most reproducible at high frequencies (CV 9–10%). Our results indicate that several methods are feasible for the determination of viscoelastic properties of bone material. The high frequency semi-dynamic test showed the highest precision within the tested nanoindentation protocols.