Search

Nondestructive cell membrane permeabilization systems enable the intracellular delivery of exogenous biomolecules for cell engineering tasks as well as the temporal sampling of cytosolic contents from live cells for the analysis of dynamic processes. Here, we report a

The emerging field of cell therapy offers the potential to treat and even cure a diverse array of diseases for which existing interventions are inadequate. Recent advances in micro and nanotechnology have added a multitude of single cell analysis methods to our research repertoire. At the same time, techniques have been developed for the precise engineering and manipulation of cells. Together, these methods have aided the understanding of disease pathophysiology, helped formulate corrective interventions at the cellular level, and expanded the spectrum of available cell therapeutic options. This review discusses how micro and nanotechnology have catalyzed the development of cell sorting, cellular engineering, and single cell analysis technologies, which have become essential workflow components in developing cell-based therapeutics. The review focuses on the technologies adopted in research studies and explores the opportunities and challenges in combining the various elements of cell engineering and single cell analysis into the next generation of integrated and automated platforms that can accelerate preclinical studies and translational research.

Manipulation of cells for applications such as biomanufacturing and cell-based therapeutics involves introducing biomolecular cargoes into cells. However, successful delivery is a function of multiple experimental factors requiring several rounds of optimization. Here, we present a high-throughput multiwell-format localized electroporation device (LEPD) assisted by deep learning image analysis that enables quick optimization of experimental factors for efficient delivery. We showcase the versatility of the LEPD platform by successfully delivering biomolecules into different types of adherent and suspension cells. We also demonstrate multicargo delivery with tight dosage distribution and precise ratiometric control. Furthermore, we used the platform to achieve functional gene knockdown in human induced pluripotent stem cells and used the deep learning framework to analyze protein expression along with changes in cell morphology. Overall, we present a workflow that enables combinatorial experiments and rapid analysis for the optimization of intracellular delivery protocols required for genetic manipulation.

Significant efforts have been devoted to enhancing the sensitivity and working range of flexible pressure sensors to improve the precise measurement of subtle variations in pressure over a wide detection spectrum. However, achieving sensitivities exceeding 1000 kPa−1 while maintaining a pressure working range over 100 kPa is still challenging because of the limited intrinsic properties of soft matrix materials. Here, we report a magnetic field-induced porous elastomer with micropillar arrays (MPAs) as sensing materials and a well-patterned nickel fabric as an electrode. The developed sensor exhibits an ultrahigh sensitivity of 10,268 kPa−1 (0.6–170 kPa) with a minimum detection pressure of 0.25 Pa and a fast response time of 3 ms because of the unique structure of the MPAs and the textured morphology of the electrode. The porous elastomer provides an extended working range of up to 500 kPa with long-time durability. The sophisticated sensor system coupled with an integrated wireless recharging system comprising a flexible supercapacitor and inductive coils for transmission achieves excellent performance. Thus, a diverse range of practical applications requiring a low-to-high pressure range sensing can be developed. Our strategy, which combines a microstructured high-performance sensor device with a wireless recharging system, provides a basis for creating next-generation flexible electronics.

Dislocation dynamic simulations are intended as a tool to understand and predict the mechanical behavior of metallic materials, but its prediction has never been directly verified by experiments due to differences in specimen strain rate and size. In this work, a comprehensive experimental framework is proposed to attempt direct comparison between experiments and discrete dislocation dynamics (DDD) modelling. By integrating high-throughput sample fabrication and a customized testing apparatus, the sample size and strain rate typically employed in DDD simulations are explored experimentally. Constitutive properties such as stress-strain response are measured, and microstructural information is obtained from transmission electron microscopy (TEM) imaging, electron backscatter diffraction (EBSD), and TEM-based orientation mapping. Magnesium and copper were selected, as case studies, to demonstrate the newly developed experimental procedure. Measured stress-strain responses for Mg are consistent with those obtained with a miniaturized Hopkison bar experiments. By exploiting the validated workflow, the effect of strain rate on micropillar heterogeneous deformation and associated dislocation plasticity were revealed. The work establishes a methodology for the systematic study of not only metals but also other materials and structures at the microscale and high strain rates.

Genome engineering of cells using CRISPR/Cas systems has opened new avenues for pharmacological screening and investigating the molecular mechanisms of disease. A critical step in many such studies, is the intracellular delivery of the gene editing machinery and the subsequent manipulation of cells. However, these workflows often involve processes such as bulk electroporation for intracellular delivery and fluorescence activated cell sorting (FACS) for cell isolation that can be harsh to sensitive cell types such as human induced pluripotent stem cells (hiPSCs). This often leads to poor viability and low overall efficacy, requiring the use of large starting samples. In this work a fully automated version of the nanofountain probe electroporation (NFP-E) system, a nanopipette-based single-cell electroporation method is presented, that provides superior cell viability and efficiency compared to traditional methods. The automated system utilizes a deep convolutional network to identify cell locations and a cell-nanopipette contact algorithm to position the nanopipette over each cell for the application of electroporation pulses. The automated NFP-E is combined with micro-confinement arrays for cell isolation to demonstrate a workflow that can be used for CRISPR/Cas9 gene editing and cell tracking with potential applications in screening studies and isogenic cell line generation.

Summary Animals' teeth have evolved to provide food procurement, mastication, and protection. These functions, directly linked to survival of many living animal species, require superior hardness and abrasion resistance in the animal's dentition system. Such resistance typically emerges from damage tolerance and sharpness preservation during the organism’s life span. An example is the sea urchin tooth, which through gradients in mechanical properties together with exploitation of microstructural features achieves such functionality. Using contact mechanics, dimensional analysis, and a novel in situ scanning electron microscopy experimental methodology, conditions for tooth deformation and wear, via a self-sharpening mechanism consisting in plate chipping, were imaged and quantified. Nonlinear finite-element modeling of the self-sharpening mechanism provided insight into the synergy between constituent material properties and tooth microstructural elements. The findings reported here should inspire the design of novel tools used in machining operations, e.g., cutting and grinding, as well as in mining and tunnel boring.

Summary During the last decade, two-dimensional (2D) materials have emerged as versatile building blocks for the next generation of engineered materials. However, the intrinsically brittle behavior of 2D materials has thus far delayed their adoption in applications such as sensors and structural materials. Herein, we demonstrate a strategy for toughening graphene oxide (GO) through synergistic interfacial interactions between GO monolayers and ultrathin layers of strongly interacting poly(vinyl alcohol) (PVA). By creating GO-PVA and PVA-GO-PVA nanolaminates, we demonstrate a 2-fold increase in GO toughness, which translates into dramatic increases in energy dissipation and piercing resistance. Atomistic simulations show that this remarkable behavior arises from a polymer chain crack-bridging mechanism, resulting from a synergistic combination of interdomain reinforcements across the GO monolayer and extensive GO-polymer interfacial hydrogen-bonding interactions. The reported findings highlight the potential for achieving engineered 2D materials with superior mechanical properties by incorporating deformation and failure-resistant mechanics arising from tailored chemical interactions between constituents.

Annihilation of vacancy clusters in monolayer molybdenum diselenide (MoSe

The fi eld of in situ nanomechanics is greatly benefi ting from microelectromechanical systems (MEMS) technology and integrated microscale testing machines that can measure a wide range of mechanical properties at nanometer scales, while characterizing the damage or microstructure evolution in electron microscopes. This article focuses on the latest advances in MEMS-based nanomechanical testing techniques that go beyond stress and strain measurements under typical monotonic loadings. Specifi cally, recent advances in MEMS testing machines now enable probing key mechanical properties of nanomaterials related to fracture, fatigue, and wear. Tensile properties can be measured without instabilities or at high strain rates, and signature parameters such as activation volume can be obtained. Opportunities for environmental in situ nanomechanics enabled by MEMS technology are also discussed.

A number of experimental techniques have been used to characterize the mechanical properties and wear of biomaterials, from nanoindentation to scratch to atomic force microscopy testing. While all these experiments provide valuable information on the mechanics and functionality of biomaterials (e.g., animals’ teeth), they lack the ability to combine the measurement of force and sliding velocities with high resolution imaging of the processes taking place at the biomaterial-substrate interface. Here we present an experiment for the in situ scanning electron microscopy characterization of the mechanics of friction and wear of biomaterials with simultaneous control of mechanical and kinematic variables. To illustrate the experimental methodology, we report the wear of the sea urchin tooth, which exhibits a unique combination of architecture and material properties tailored to withstand abrasion loads in different directions. By quantifying contact conditions and changes in the tooth tip geometry, we show that the developed methodology provides a versatile and promising tool to investigate wear mechanisms in a variety of animal teeth accounting for microscale effects.

Nanomechanical resonators make exquisite force sensors due to their small footprint, low dissipation, and high frequencies. Because the lowest resolvable force is limited by ambient thermal noise, resonators are either operated at cryogenic temperatures or coupled to a high-finesse optical or microwave cavity to reach sub aN Hz-1/2 sensitivity. Here, we show that operating a monolayer WS2 nanoresonator in the strongly nonlinear regime can lead to comparable force sensitivities at room temperature. Cavity interferometry was used to transduce the nonlinear response of the nanoresonator, which was characterized by multiple pairs of 1:1 internal resonance. Some of the modes exhibited exotic line shapes due to the appearance of Hopf bifurcations, where the bifurcation frequency varied linearly with the driving force and forms the basis of the advanced sensing modality. The modality is less sensitive to the measurement bandwidth, limited only by the intrinsic frequency fluctuations, and therefore, advantageous in the detection of weak incoherent forces.

Electrospun carbon nanofibers, produced from polyacrylonitrile (PAN) nanofiber precursors, with their superior mechanical properties, are promising candidates for manufacturing advanced polymer composites. Here, we report a series of tensile tests performed in situ under scanning electron microscope (SEM)/transmission electron microscope (TEM) observation, which show that the modulus and strength of electrospun carbon nanofibers can be enhanced through a simple mechanical constraint during the carbonization step in the electrospinning process. The constrained carbon nanofibers of diameter less than 150 nm were nanomanipulated inside the SEM onto a specialized microelectromechanical systems (MEMS) based testing platform and subsequently tested in uniaxial tension until failure. It was identified that both the strength and modulus of the constrained carbon nanofibers with sub-150 nm diameters are on average higher compared to their unconstrained counterparts by ∼22% and ∼31% respectively. Also, by evaluating the internal graphitic order of the constrained carbon nanofibers using TEM-based diffraction methods, we identified that the mechanical constraint during carbonization results in a better degree of orientation in the graphitic crystallites along the fiber axis. Finally, we use Weibull statistics for the deconvolution of the effects of diameter and the mechanical constraint on the tensile properties of carbon nanofibers. The Weibull analysis also showed that the comparatively superior strength of CCNFs is primarily due to better alignment of crystallites with the fiber axis.

Nanoelectromechanical (NEMS) systems fabricated using atomically thin materials have low mass and high stiffness and are thus ideal candidates for force and mass sensing applications. Transition metal dichalcogenides (TMDCs) offer certain unique properties in their few-layered form – such as piezoelectricity and a direct band gap (in some cases) – and are an interesting alternative to graphene based NEMS. Among the demonstrated methods for displacement transduction in NEMS, cavity-interferometry provides exquisite displacement sensitivity. Typically, interferometric measurements are complemented with Raman spectroscopy to characterize the number of layers in 2D materials, and the measurements necessitate high vacuum conditions to eliminate viscous damping. Here, we report an experimental setup that facilitates both Raman spectroscopy and interferometric measurements on few-layered Tungsten Disulfide (WS2) resonators in high vacuum (

Single-cell delivery platforms like microinjection and nanoprobe electroporation enable unparalleled control over cell manipulation tasks but are generally limited in throughput. Here, we present an automated single-cell electroporation system capable of automatically detecting cells with artificial intelligence (AI) software and delivering exogenous cargoes of different sizes with uniform dosage. We implemented a fully convolutional network (FCN) architecture to precisely locate the nuclei and cytosol of six cell types with various shapes and sizes, using phase contrast microscopy. Nuclear staining or reporter fluorescence was used along with phase contrast images of cells within the same field of view to facilitate the manual annotation process. Furthermore, we leveraged the near-human inference capabilities of the FCN network in detecting stained nuclei to automatically generate ground-truth labels of thousands of cells within seconds, and observed no statistically significant difference in performance compared to training with manual annotations. The average detection sensitivity and precision of the FCN network were 95±1.7% and 90±1.8%, respectively, outperforming a traditional image-processing algorithm (72±7.2% and 72±5.5%) used for comparison. To test the platform, we delivered fluorescent-labeled proteins into adhered cells and measured a delivery efficiency of 90%. As a demonstration, we used the automated single-cell electroporation platform to deliver Cas9-guide RNA (gRNA) complexes into an induced pluripotent stem cell (iPSC) line to knock out a green fluorescent protein-encoding gene in a population of ~200 cells. The results demonstrate that automated single-cell delivery is a useful cell manipulation tool for applications that demand throughput, control, and precision.

Nature's masterfully synthesized biological materials take on greater relevance when viewed through the perspective of evolutionary abundance. The fact that beetles (order Coleoptera) account for a quarter of all extant lifeforms on Earth, makes them prime exponents of evolutionary success. In fact, their forewings are acknowledged as key traits to their radiative-adaptive success, which makes the beetle elytra a model structure for next-generation bioinspired synthetic materials. In this work, the multiscale morphological and mechanical characteristics of a variety of beetle species from the Cetoniinae subfamily are investigated with the aim of unraveling the underlying principles behind Nature's adaptation of the elytral bauplan to differences in body weight spanning three orders of magnitude. Commensurate with the integral implications of size variation in organisms, a combined material, morphological, and mechanical characterization framework, across spatial scales, was pursued. The investigation revealed the simultaneous presence of size-invariant strategies (chemical compositions, layered-fibrous architectures, graded motifs) as well as size-dependent features (scaling of elytral layers and characteristic dimensions of building blocks), synergistically combined to achieve similar levels of biomechanical functionality (stiffness, energy absorption, strength, deformation and toughening mechanisms) in response to developmental and selection constraints. The integral approach here presented seeks to shed light on Nature's solution to the problem of size variation, which underpins the diversity of beetles and the living world.

In vitro and ex vivo intracellular delivery methods hold the key for releasing the full potential of tissue engineering, drug development, and many other applications. In recent years, there has been significant progress in the design and implementation of intracellular delivery systems capable of delivery at the same scale as viral transfection and bulk electroporation but offering fewer adverse outcomes. This review strives to examine a variety of methods for in vitro and ex vivo intracellular delivery such as flow-through microfluidics, engineered substrates, and automated probe-based systems from the perspective of throughput and control. Special attention is paid to a particularly promising method of electroporation using micro/nanochannel based porous substrates, which expose small patches of cell membrane to permeabilizing electric field. Porous substrate electroporation parameters discussed include system design, cells and cargos used, transfection efficiency and cell viability, and the electric field and its effects on molecular transport. The review concludes with discussion of potential new innovations which can arise from specific aspects of porous substrate-based electroporation platforms and high throughput, high control methods in general.

Kirigami structures provide a promising approach to transform flat films into 3D complex structures that are difficult to achieve by conventional fabrication approaches. By designing the cutting geometry, it is shown that distinct buckling-induced out-of-plane configurations can be obtained, separated by a sharp transition characterized by a critical geometric dimension of the structures. In situ electron microscopy experiments reveal the effect of the ratio between the in-plane cut size and film thickness on out-of-plane configurations. Moreover, geometrically nonlinear finite element analyses (FEA) accurately predict the out-of-plane modes measured experimentally, their transition as a function of cut geometry, and provide the stress-strain response of the kirigami structures. The combined computational-experimental approach and results reported here represent a step forward in the characterization of thin films experiencing buckling-induced out-of-plane shape transformations and provide a path to control 3D configurations of micro- and nanoscale buckling-induced kirigami structures. The out-of-plane configurations promise great utility in the creation of micro- and nanoscale systems that can harness such structural behavior, such as optical scanning micromirrors, novel actuators, and nanorobotics. This work is of particular significance as the kirigami dimensions approach the sub-micrometer scale which is challenging to achieve with conventional micro-electromechanical system technologies.

Annihilation of vacancy clusters in monolayer molybdenum diselenide (MoSe2 ) under electron beam irradiation is reported. In situ high-resolution transmission electron microscopy observation reveals that the annihilation is achieved by diffusion of vacancies to the free edge near the vacancy clusters. Monte Carlo simulations confirm that it is energetically favorable for the vacancies to locate at the free edge. By computing the minimum energy path for the annihilation of one vacancy cluster as a case study, it is further shown that electron beam irradiation and pre-stress in the suspended MoSe2 monolayer are necessary for the vacancies to overcome the energy barriers for diffusion. The findings suggest a new mechanism of vacancy healing in 2D materials and broaden the capability of electron beam for defect engineering of 2D materials, a promising way of tuning their properties for engineering applications.

Combining experimental and computational studies of nanocomposite interfaces is highly needed to gain insight into their performance. However, there are very few literature reports, combining well-controlled atomic force microscopy experiments with molecular dynamic simulations, which explore the role of polymer chemistry and assembly on interface adhesion and shear strength. In this work, we investigate graphene oxide (GO)-polymer interfaces prevalent in nanocomposites based on a nacre-like architectures. We examine the interfacial strength resulting from van der Waals and hydrogen bonding interactions by comparing the out-of-plane separation and in-plane shear deformations of GO-polyethylene glycol (PEG) and GO-polyvinyl alcohol (PVA). The investigation reveals an overall better mechanical performance for the anhydrous GO-PVA system in both out-of-plane and in-plane deformation modes, highlighting the benefits of the donor-acceptor hydrogen bond formation present in GO-PVA. Such bond formation results in inter-chain hydrogen bond networks leading to stronger interfaces. By contrast, PEG, a hydrogen bond acceptor only, relies primarily on van der Waals inter-chain interactions, typically resulting in weaker interactions. The study also predicts that water addition increases the adhesion of GO-PEG but decreases the adhesion of GO-PVA, and slightly increases the shear strength in both systems. Furthermore, by comparing simulations and experiments, we show that the CHARMM force field has enough accuracy to capture the effect of polymer content, water distribution, and to provide quantitative guidance for achieving optimum interfacial properties. Therefore, the study demonstrates an effective methodology, in the Materials Genome spirit, toward the design of 2D materials-polymer nanocomposites system for applications demanding mechanical robustness.

Measuring changes in enzymatic activity over time from small numbers of cells remains a significant technical challenge. In this work, a method for sampling the cytoplasm of cells is introduced to extract enzymes and measure their activity at multiple time points. A microfluidic device, termed the live cell analysis device (LCAD), is designed, where cells are cultured in microwell arrays fabricated on polymer membranes containing nanochannels. Localized electroporation of the cells opens transient pores in the cell membrane at the interface with the nanochannels, enabling extraction of enzymes into nanoliter-volume chambers. In the extraction chambers, the enzymes modify immobilized substrates, and their activity is quantified by self-assembled monolayers for matrix-assisted laser desorption/ionization (SAMDI) mass spectrometry. By employing the LCAD-SAMDI platform, protein delivery into cells is demonstrated. Next, it is shown that enzymes can be extracted, and their activity measured without a loss in viability. Lastly, cells are sampled at multiple time points to study changes in phosphatase activity in response to oxidation by hydrogen peroxide. With this unique sampling device and label-free assay format, the LCAD with SAMDI enables a powerful new method for monitoring the dynamics of cellular activity from small populations of cells.

Introducing exogenous molecules into cells with high efficiency and dosage control is a crucial step in basic research as well as clinical applications. Here, the capability of the nanofountain probe electroporation (NFP-E) system to deliver proteins and plasmids in a variety of continuous and primary cell types with appropriate dosage control is reported. It is shown that the NFP-E can achieve fine control over the relative expression of two cotransfected plasmids. Finally, the dynamics of electropore closure after the pulsing ends with the NFP-E is investigated. Localized electroporation has recently been utilized to demonstrate the converse process of delivery (sampling), in which a small volume of the cytosol is retrieved during electroporation without causing cell lysis. Single-cell temporal sampling confers the benefit of monitoring the same cell over time and can provide valuable insights into the mechanisms underlying processes such as stem cell differentiation and disease progression. NFP-E parameters that maximize the membrane resealing time, which is essential for increasing the sampled volume and in meeting the challenge of monitoring low copy number biomarkers, are identified. Its application in CRISPR/Cas9 gene editing, stem cell reprogramming, and single-cell sampling studies is envisioned.

Cells and tissues sense, respond to and translate mechanical forces into biochemical signals through mechanotransduction, which governs individual cell responses that drive gene expression, metabolic pathways and cell motility, and determines how cells work together in tissues. Mechanotransduction often depends on cytoskeletal networks and their attachment sites that physically couple cells to each other and to the extracellular matrix. One way that cells associate with each other is through Ca2+-dependent adhesion molecules called cadherins, which mediate cell–cell interactions through adherens junctions, thereby anchoring and organizing the cortical actin cytoskeleton. This actin-based network confers dynamic properties to cell sheets and developing organisms. However, these contractile networks do not work alone but in concert with other cytoarchitectural elements, including a diverse network of intermediate filaments. This Review takes a close look at the intermediate filament network and its associated intercellular junctions, desmosomes. We provide evidence that this system not only ensures tissue integrity, but also cooperates with other networks to create more complex tissues with emerging properties in sensing and responding to increasingly stressful environments. We will also draw attention to how defects in intermediate filament and desmosome networks result in both chronic and acquired diseases.

Naturally occurring biological materials with stiff fibers embedded in a ductile matrix are commonly known to achieve excellent balance between stiffness, strength and ductility. In particular, biological composite materials with helicoidal architecture have been shown to exhibit enhanced damage tolerance and increased impact energy absorption. However, the role of fiber reorientation inside the flexible matrix of helicoid composites on their mechanical behaviors have not yet been extensively investigated. In the present work, we introduce a Discontinuous Fiber Helicoid (DFH) composite inspired by both the helicoid microstructure in the cuticle of mantis shrimp and the nacreous architecture of the red abalone shell. We employ 3D printed specimens, analytical models and finite element models to analyze and quantify in-plane fiber reorientation in helicoid architectures with different geometrical features. We also introduce additional architectures, i.e., single unidirectional lamina and mono-balanced architectures, for comparison purposes. Compared with associated mono-balanced architectures, helicoid architectures exhibit less fiber reorientation values and lower values of strain stiffening. The explanation for this difference is addressed in terms of the measured in-plane deformation, due to uniaxial tensile of the laminae, correlated to lamina misorientation with respect to the loading direction and lay-up sequence.

Graphene oxide (GO), whose highly tunable surface chemistry enables the formation of strong interfacial hydrogen-bond networks, has garnered increasing interest in the design of devices that operate in the presence of water. For instance, previous studies have suggested that controlling GO's surface chemistry leads to enhancements in interfacial shear strength, allowing engineers to manage deformation pathways and control failure mechanisms. However, these previous reports have not explored the role of ambient humidity and only offer extensive chemical modifications to GO's surface as the main pathway to control GO's interfacial properties. Herein, through atomic force microscopy experiments on GO-GO interfaces, the adhesion energy and interfacial shear strength of GO were measured as a function of ambient humidity. Experimental evidence shows that adhesion energy and interfacial shear strength can be improved by a factor of 2-3 when GO is exposed to moderate (∼30% water weight) water content. Furthermore, complementary molecular dynamics simulations uncovered the mechanisms by which these nanomaterial interfaces achieve their properties. They reveal that the strengthening mechanism arises from the formation of strongly interacting hydrogen-bond networks, driven by the chemistry of the GO basal plane and intercalated water molecules between two GO surfaces. In summary, the methodology and findings here reported provide pathways to simultaneously optimize GO's interfacial and in-plane mechanical properties, by tailoring the chemistry of GO and accounting for water content, in engineering applications such as sensors, filtration membranes, wearable electronics, and structural materials.

In the field of genetic engineering, the modification of genes to produce stable cell lines has a variety of applications ranging from the development of novel therapeutics to patient specific treatments. To successfully generate a cell line, the gene of interest must be delivered into the cell and integrated into the genome. The efficiency of cell line generation systems therefore depends on the efficiency of delivery of genetically modifying molecules such as plasmids and CRISPR/CAS9 complexes. In this work, we describe a localized electroporation-based system to generate stable monoclonal cell lines. By employing the nanofountain probe electroporation (NFP-E) system, single cells in patterned cultures are selectively transfected with plasmids, grown, and harvested to obtain stably expressing cell lines. Methods for microcontact printing, cell culture, electroporation, and harvesting are detailed in this chapter.

Bighorn sheep ( Ovis canadensis ) rams hurl themselves at each other at speeds of ∼9 m/s (20 mph) to fight for dominance and mating rights. This necessitates impact resistance and energy absorption mechanisms, which stem from material-structure components in horns. In this study, the material hierarchical structure as well as correlations between the structure and mechanical properties are investigated. The major microstructural elements of horns are found as tubules and cell lamellae, which are oriented with (∼30⁰) angle with respect to each other. The cell lamellae contain keratin cells, in the shape of pancakes, possessing an average thickness of ∼2 µm and diameter of ∼20–30 µm. The morphology of keratin cells reveals the presence of keratin fibers and intermediate filaments with diameter of ∼200 nm and ∼12 nm, respectively, parallel to the cell surface. Quasi-static and high strain rate impact experiments, in different loading directions and hydration states, revealed a strong strain rate dependency for both dried and hydrated conditions. A strong anisotropy behavior was observed under impact for the dried state. The results show that the radial direction is the most preferable impact orientation because of its superior energy absorption. Detailed failure mechanisms under the aforementioned conditions are examined by bar impact recovery experiments. Shear banding, buckling of cell lamellae, and delamination in longitudinal and transverse direction were identified as the cause for strain softening under high strain rate impact. While collapse of tubules occurs in both quasi-static and impact tests, in radial and transverse directions, the former leads to more energy absorption and impact resistance. Statement of Significance Bighorn sheep ( Ovis canadensis ) horns show remarkable impact resistance and energy absorption when undergoing high speed impact during the intraspecific fights. The present work illustrates the hierarchical structure of bighorn sheep horn at different length scales and investigates the energy dissipation mechanisms under different strain rates, loading orientations and hydration states. These results demonstrate how horn dissipates large amounts of energy, thus provide a new path to fabricate energy absorbent and crashworthiness engineering materials.

Desmoplakin connects desmosomal core components to intermediate filaments at sites of cell–cell adhesion. Modulating the strength of this linkage using desmoplakin mutants led to alterations in cell–substrate and cell–cell forces and cell stiffness as assessed by micropillar arrays and atomic force microscopy. Perturbation of the actin cytoskeleton leads to abrogation of these effects., The translation of mechanical forces into biochemical signals plays a central role in guiding normal physiological processes during tissue development and homeostasis. Interfering with this process contributes to cardiovascular disease, cancer progression, and inherited disorders. The actin-based cytoskeleton and its associated adherens junctions are well-established contributors to mechanosensing and transduction machinery; however, the role of the desmosome–intermediate filament (DSM–IF) network is poorly understood in this context. Because a force balance among different cytoskeletal systems is important to maintain normal tissue function, knowing the relative contributions of these structurally integrated systems to cell mechanics is critical. Here we modulated the interaction between DSMs and IFs using mutant forms of desmoplakin, the protein bridging these structures. Using micropillar arrays and atomic force microscopy, we demonstrate that strengthening the DSM–IF interaction increases cell–substrate and cell–cell forces and cell stiffness both in cell pairs and sheets of cells. In contrast, disrupting the interaction leads to a decrease in these forces. These alterations in cell mechanics are abrogated when the actin cytoskeleton is dismantled. These data suggest that the tissue-specific variability in DSM–IF network composition provides an opportunity to differentially regulate tissue mechanics by balancing and tuning forces among cytoskeletal systems.

Flight is not the exclusive domain of birds; mammals (bats), insects, and some fish have independently developed this ability by the process of convergent evolution. Birds, however, greatly outperform other flying animals in efficiency and duration; for example the common swift (Apus apus) has recently been reported to regularly fly for periods of 10 months during migration. Birds owe this extraordinary capability to feathers and bones, which are extreme lightweight biological materials. They achieve this crucial function through their efficient design spanning multiple length scales. Both feathers and bones have unusual combinations of structural features organized hierarchically from nano- to macroscale and enable a balance between lightweight and bending/torsional stiffness and strength. The complementary features between the avian bone and feather are reviewed here, for the first time, and provide insights into nature's approach at creating structures optimized for flight. We reveal a novel aspect of the feather vane, showing that its barbule spacing is consistently within the range 8–16 μm for birds of hugely different masses such as Anna's Hummingbird (Calypte anna) (4 g) and the Andean Condor (Vultur gryphus) (11,000 g). Features of the feather and bone are examined using the structure-property relationships that define Materials Science. We elucidate the role of aerodynamic loading on observed reinforced macrostructural features and efficiently tailored shapes adapted for specialized applications, as well as composite material utilization. These unique features will inspire synthetic structures with maximized performance/weight for potential use in future transportation systems.

Kirigami with a variety of cut patterns have been recently investigated as means to transform 2D surfaces into 3D structures, offering a number of functionalities. In this work, we show that a single Kirigami motif, defining two inner panels connected by hinges, can generate a rich variety of out-of-plane symmetric and asymmetric deformation modes. The out-of-plane responses, caused by local instabilities when a far field tensile load is applied, manifest themselves at the inner plates, exhibiting a combination of one- and two-dimensional rotations (tilt and twist), effectively morphing the planar geometries into complex 3D surfaces. By conducting numerical analyses and experiments, the relationship between changes in the cut geometry, and out-of-plane responses is identified. In particular, two types of instabilities emerge, coincident bifurcation points for both inner plates, responsible for two-dimensional tilts, and sequential bifurcation points, which produce one-dimensional tilts with distinct buckling thresholds. Nonlinear parametric finite element analysis (FEA) is used to explore the space of possible configurations and characterize the behavior of the structure as a function of its parameters, revealing a variety of deformation modes and structural responses of interest. Numerical predictions are experimentally validated using paper Kirigami and the shadow Moire technique. The combined numerical–experimental approach reveals a plethora of programmable and controllable deformation modes with intrinsic nonlinear behaviors, enabled by tunable imperfections, which promise to influence a variety of applications ranging from light modulation to fluid flow control.

Graphene oxide (GO) shows promise as a nanocomposite building block due to its exceptional mechanical properties. While atomistic simulations have become central to investigating its mechanical properties, the method remains prohibitively expensive for large deformations and mesoscale failure mechanisms. To overcome this, we establish a coarse-grained (CG) model that captures key mechanical and interfacial properties, and the non-homogeneous effect of oxidation in GO sheets. The CG model consists of three types of CG beads, representing groups of pristine sp2 carbon atoms, and hydroxyl and epoxide functionalized regions. The CG force field is parameterized based on density functional-based tight binding simulations on three extreme cases. It accurately quantifies deterioration of tensile modulus and strength at the expense of improving interlayer adhesion with increasing oxidation of varying chemical compositions. We demonstrate the applicability of the model to study mesoscale phenomena by reproducing different force vs. indentation curves in silico, corroborating recent experimental observations on how chemistry near contact point influences properties. Finally, we apply the model to measure the fracture toughness of pristine graphene and GO. The critical stress intensity factor ( K c ) of graphene is found to be the highest, and epoxide-rich GO also possesses higher K c compared to hydroxyl-rich GO.

Several recent micro- and nanotechnologies have provided novel methods for biological studies of adherent cells because the small features of these new biotools provide unique capabilities for accessing cells without the need for suspension or lysis. These novel approaches have enabled gentle but effective delivery of molecules into specific adhered target cells, with unprecedented spatial resolution. We review here recent progress in the development of these technologies with an emphasis on in vitro delivery into adherent cells utilizing mechanical penetration or electroporation. We discuss the major advantages and limitations of these approaches and propose possible strategies for improvements. Finally, we discuss the impact of these technologies on biological research concerning cell-specific temporal studies, for example non-destructive sampling and analysis of intracellular molecules.

In PNAS, Cao et al. (1) employ track-etched membranes for the intracellular delivery of various molecular cargoes via electroporation in both adherent and suspended cells. While the concept of utilizing such membranes for electroporation is not novel (2, 3), its extension to suspended cells and the wide parametric space covered in the article are noteworthy. The objectives of this letter are to correct a minor lapse in the article’s introduction, as well as to put the results in a broader context. In ref. 1, the authors cite that the delivery efficiency of a plasmid, using track-etched membranes and localized electroporation (2), was less than 50% (reference 16 in ref. 1). However, an efficiency … [↵][1]1To whom correspondence may be addressed. Email: espinosa{at}northwestern.edu. [1]: #xref-corresp-1-1

This paper describes a systematic study on the nanoscale toughening of monolayer graphene oxide (GO) by an ultra-thin polymer adlayer, which impedes the propagation of cracks during intraplanar fracture. Using molecular dynamics simulations, the crack-bridging capabilities of a library of five hydrogen-bonding-capable polymers are explored against an epoxide-rich GO substrate. The best crack-bridging effect is found in polymers with functional groups that can both donate/accept hydrogen atoms and have better capability to form cooperative hydrogen bonds. Aligning the chains of poly(acrylic acid) orthogonally to the crack propagation direction significantly enhances the fracture toughness of monolayer GO (by 310%) in comparison to that for an adlayer with randomly arranged chains (180% enhancement). Notably, van der Waals interactions, which are seldom highlighted in the fabrication of strong GO-polymer interfaces, are found to also provide significant crack-bridging capabilities when the polymers possess large side groups. These results pave the way for a set of design criteria that can help in remediating the intrinsically brittle mechanical behavior of two-dimensional materials, a barrier that currently restricts their potential applications.

Nanomechanical resonators make exquisite force sensors due to their small footprint, low dissipation, and high frequencies. Because the lowest resolvable force is limited by ambient thermal noise, resonators are either operated at cryogenic temperatures or coupled to a high-finesse optical or microwave cavity to reach sub aN Hz