Intact Material Research Articles

Cannabidiol (CBD) Oil Does Not Display an Entourage Effect in Reducing Cancer Cell Viability in vitro

Introduction: Several studies have found that cannabinoids, particularly delta-9-tetrahydrocannabinol and cannabidiol (CBD), have the ability to reduce cancer cell viability. An ongoing debate regarding the use of medical Cannabis revolves around the effectiveness of pure compounds versus intact plant material for treatment. Proponents for the use of intact plant material or botanical extracts argue that there is a synergistic effect between the different cannabinoids, terpenoids, and flavonoids; this is commonly referred to as the “entourage effect.” Our study was designed to test the validity of the proposed entourage effect in a narrow application using a cancer cell viability model. Materials and Methods: Six cancer cell lines, from 3 different types of human cancer were treated with 10 μM pure CBD or 10 μM CBD from hemp (Cannabis sativa) oil (obtained from 3 different commercial sources) for 48 h, and cell viability was measured with the MTS assay. Dose-response curves were then performed to compare the potencies of pure CBD to CBD oils. CBD concentrations were independently confirmed in the commercial oils, and cannabinoid and terpene composition were also compared. Results: CBD (10 μM) was able to reduce cell viability in 3 of the 6 cell lines tested, and this was found to be cell line specific and not specific to select cancers. None of the CBD oils tested were able to reduce viability to a greater extent than that of pure CBD. Additionally, dose-response curves found lower IC₅₀ values for pure CBD compared to the most potent CBD oil tested. Interestingly, some oils actually appeared to protect cancer cells from the effects of CBD. Conclusions: We found that pure CBD was as potent or more potent at reducing cancer cell viability as the most potent oil tested, suggesting that there is no “entourage” effect under these specific in vitro conditions.

A two‐scale constitutive framework for modelling localised deformation in saturated dilative hardening material

SummaryUndrained deformation of dilative sand generates negative excess pore pressure. It enhances the strength, which is called dilative hardening. This increased suction is not permanent. The heterogeneity at the grain scale triggers localisations causing local volume changes. The negative hydraulic gradient drives fluid into dilating shear zones. It loosens the soil and diminishes the shear strength. It is essential to understand the mechanism behind this internal drainage and to capture it numerically. The purpose of this paper is to develop a macroscopic constitutive relationship for the undrained deformation of saturated dense sand in the presence of a locally fully or partially drained shear band. Separate constitutive relations are generated for the band and intact material. Both time and scale dependence during pore fluid diffusion in saturated sand are captured, eliminating the mesh dependency for finite element implementations. The model is applied to the Gauss points that satisfy the bifurcation criterion. The proposed method is calibrated to recreate the undrained macroscopic response bestowed by an extra‐small mesh. The microscopic behaviours inside and outside shear band predicted by this model are qualitatively in good agreement with individual material point behaviours inside and outside the shear band in the extra‐small mesh. Depending on the loading rate and the shear band thickness, the response inside the band can be fully or partially drained, which governs the ultimate global strength. The calibrated model is exploited to simulate an upscaled biaxial compression test with semipermeable boundaries.

Robust detection of oncometabolic aberrations by 1H\u201313C heteronuclear single quantum correlation in intact biological specimens

Magnetic resonance (MR) spectroscopy has potential to non-invasively detect metabolites of diagnostic significance for precision oncology. Yet, many metabolites have similar chemical shifts, yielding highly convoluted 1H spectra of intact biological material and limiting diagnostic utility. Here, we show that hydrogen–carbon heteronuclear single quantum correlation (1H–13C HSQC) offers dramatic improvements in sensitivity compared to one-dimensional (1D) 13C NMR and significant signal deconvolution compared to 1D 1H spectra in intact biological settings. Using a standard NMR spectroscope with a cryoprobe but without specialized signal enhancing features such as magic angle spinning, metabolite extractions or 13C-isotopic enrichment, we obtain well-resolved 2D 1H–13C HSQC spectra in live cancer cells, in ex vivo freshly dissected xenografted tumors and resected primary tumors. This method can identify tumors with specific oncometabolite alterations such as IDH mutations by 2-hydroxyglutarate and PGD-deleted tumors by gluconate. Results suggest potential of 1H–13C HSQC as a non-invasive diagnostic in precision oncology.

Oxidative Stress in Male Infertility: Causes, Effects in Assisted Reproductive Techniques, and Protective Support of Antioxidants.

The spermatozoon is a highly specialized cell, whose main function is the transport of the intact male genetic material into the oocyte. During its formation and transit throughout male and female reproductive tracts, sperm cells are internally and externally surrounded by reactive oxygen species (ROS), which are produced from both endogenous and exogenous sources. While low amounts of ROS are known to be necessary for crucial physiological sperm processes, such as acrosome reaction and sperm–oocyte interaction, high levels of those species underlie misbalanced antioxidant-oxidant molecules, generating oxidative stress (OS), which is one of the most damaging factors that affect sperm function and lower male fertility potential. The present work starts by reviewing the different sources of oxidative stress that affect sperm cells, continues by summarizing the detrimental effects of OS on the male germline, and discusses previous studies addressing the consequences of these detrimental effects on natural pregnancy and assisted reproductive techniques effectiveness. The last section is focused on how antioxidants can counteract the effects of ROS and how sperm fertilizing ability may benefit from these agents.

Experimental study on the strength, deformation and crack evolution behaviour of red sandstone samples containing two ice-filled fissures under triaxial compression

The artificial freezing method is an effective method of shaft construction in water-rich soft rock beds. Engineering frozen walls involves the use of intact rock material, flaws and ice. To better understand the mechanical properties of frozen walls, explore the failure laws of frozen fractured rock, and solve the problem of complex ice-rock coupling, it is necessary to carry out laboratory triaxial tests on frozen fractured rock. Herein, X-ray diffraction and mesostructure observations are used to analyse the mineral composition and microstructural characteristics of red sandstone specimens taken from the Shilawusu coal mine. Then, the strength, deformation and crack evolution behaviour of red sandstone containing two pre-existing ice-filled fissures are obtained under triaxial compression using the self-developed DRTS−500 subzero rock triaxial testing system. The experimental results show that the peak strength and elastic modulus of the specimens with pre-existing flaws are all much lower than those of the intact specimens, and the strength parameters are distinctly related to the flaw combinations. For the frozen red sandstone containing two intermittent ice-filled fissures, the peak strength, elastic modulus and shear strength parameters all have a linear relationship with the confining pressure and temperature, whereas the shear strength (σ1 − σ3) has a nonlinear relationship with the confining pressure. The deviatoric stress-strain curve of the samples is divided by four characteristic stresses (compacting stress, crack initiation stress, dilatancy stress and peak stress) into five stages from loading to failure. The lower the confining pressure or temperature is, the faster the curve falls after reaching the stress-strain peak value, indicating that the sample exhibits increasingly brittle characteristics. The ultimate failure modes of the specimens containing two pre-existing ice-filled flaws include mixtures of several types of cracks (shear cracks, main cracks, secondary cracks, tensile cracks, coplanar cracks, slip cracks and wing cracks), which depend on the flaw combinations. An obvious crack coalescence failure mode can be observed in the GG and GS specimens, but no obvious crack coalescence phenomenon can be observed in the SS specimens.

Is a Partial Anterior Cruciate Ligament Tear Truly Partial? A Clinical, Arthroscopic, and Histologic Investigation

(1) To determine whether examination under anesthesia (EUA) and/or magnetic resonance imaging (MRI) is an accurate means for determining patient selection for a selective-bundle reconstruction, (2) to determine whether the preoperative clinical assessment correlates with the arthroscopic examination, and (3) to examine histologically whether a partial anterior cruciate ligament (ACL) tear is truly partial in terms of vascular insult. This prospective, nonrandomized cohort study included 95 patients who underwent primary ACL reconstruction from January 2013 through May 2014. All patients underwent an EUA, MRI, and arthroscopic evaluation. In patients with partial ACL tears, the intact bundle was resected and underwent histologic examination. The χ2 test was used to compare EUA and MRI in the detection of partial tears. Of the 95 patients included, 40 (42%) had EUA findings consistent with a partial ACL tear whereas 22 (23%) had MRI findings interpreted as showing a partial ACL tear. Arthroscopic examination confirmed a partial ACL tear in only 11 patients (12%). The sensitivity of EUA and MRI in the detection of partial ACL tears was 100% and 90.9%, and the specificity was 65% and 85.7%, respectively. The χ2 test suggested statistically significant associations between the method of evaluation and diagnostic outcome [χ2(1)= 7.83, P= .005]. MRI was 1.24 times more likely to correctly identify a partial tear (95% confidence interval, 1.06-1.45). EUA was 2.23 times as likely to incorrectly identify a partial tear (95% confidence interval, 1.24-4.02). The histologic analysis showed increased numbers of lymphocytes, absent polymorphonuclear leukocytes, predominant fibroblasts, neovascularization, and variable collagen orientation. There is a disparity between EUA, MRI, and arthroscopic findings in the evaluation of partial ACL tears. Arthroscopy remains the gold standard for diagnosing the macroscopic integrity of the intact bundle. Microscopic analysis reveals that the integrity of the remaining intact ligament material is altered and may show a histologic response similar to a complete ACL rupture. Level III, prospective, nonrandomized cohort study.

Rapid, non-destructive analysis of calcium and strontium in eggshells by WD-XRF

We developed a simple method of analyzing the strontium (Sr) and calcium (Ca) content of intact eggshell samples in support of a broader study of how dietary Sr uptake impacts waterfowl eggshell quality. We used wavelength dispersive - x-ray fluorescence spectrometry (WD-XRF) to analyze eggshell pieces ranging in size from ∼6-mm2 fragments to intact half-shells. We verified this approach on a subset of reference shells by subjecting the same region and volume of shell material from which x-ray signals were measured to analysis by inductively-coupled plasma mass spectrometry (ICP-MS). An analysis of the sources of analytical uncertainty yielded total internal error estimates of ±0.3 and 5% relative for Ca and Sr, respectively, on the basis of which the chemistry of intact shell material analyzed by WD-XRF in this study is compared. The total external errors associated with the WD-XRF results of this study in relation to certified reference material (National Institute of Standards and Technology [NIST] 1400 [a bone ash]) are ±9 and 13.5% relative for Ca and Sr, respectfully (95% CL). Our results demonstrate this method is acceptably accurate and precise for many wildlife management applications. WD-XRF analysis is a quick and inexpensive alternative to traditional methods for determining eggshell Sr and Ca that require acid digestion, allowing for generation of larger datasets that might otherwise be cost-prohibitive, while preserving sample material intact.

Kinetics of healing of asphalt mixtures

Abstract Promotion of healing of asphalt mixtures is significant to enhance the resiliency and extend the service life of asphalt pavements. Extensive laboratory measurements of effects of healing and field observations have proven its impact. Many approaches have been developed to explain and model healing of asphalt mixtures based on its mechanisms and/or mechanical features. However, due to the complexity caused by the co-existence of several coordinated rate processes during healing, there are still unclear aspects in terms of mechanisms and appropriate representation of each rate process. Target this issue, this study proposes to investigate healing of asphalt mixtures using kinetic and thermodynamic quantities and formulations, aiming at clarifying the relationships among the coordinated processes involved in healing and developing an approach to predict the rate of each process. Healing is found to contain two coordinated rate processes: free energy change and bond restoration. Healing rate is proportional to the rates of these two processes, and the healing activation energy is the product of the activation energy of the free energy change and that of the bond restoration. In order to calculate each activation energy and rate constant, the recoverable strain energy (RSE) and recoverable pseudo strain energy (RPSE) measured in the rest time of a creep and step-loading recovery (CSR) test are used. The RPSE in the intact material and the surface energy constitute the total free energy change for healing. A three-parameter model is proposed to simulate a rate process like the change of the RSE or free energy change with time, and a dimensionless logarithmic rate is defined to obtain a representative change rate. This representative rate is ideal to be used as the rate constant in the Arrhenius equation to compute the activation energy. The activation energies for viscoelastic recovery, healing, free energy change, and bond restoration are determined for two types of asphalt mixtures at unaged and aged states. The relations between different activation energies are clarified so as the relationships among different rate processes. Furthermore, the healing activation energy obtained from this kinetic approach is proven to be the same as that calculated using the damage density from a mechanistic approach. A method to generate a damage density-based healing curve at any rest time at any temperature using just energy measurements is proposed in this study.

Numerical modeling of geomaterial fracture using a cohesive crack model in grain-based DEM

This paper aims to study the mechanical behavior of rocks using the cohesive crack model (CCM) and grain-based model (GBM) within the distinct element method (DEM) simulations. In the GBM-DEM, the Voronoi tessellation scheme is used and the intact material is simulated as assemblies of a number of particles bonded together at their contact areas. Implemented CCM defines the nonlinear behavior of the grain interfaces under various modes of loading. Numerical simulation of a tension–compression and direct shear tests is conducted to verify the correct implementation of CCM. Moreover, in the application of such a numerical framework, the uniaxial and biaxial compression tests as well as the Brazilian disc test are simulated in the universal distinct element code using a calibration process and then the results are compared with the relevant responses of the rock sample. Finally, the capability of soft computing method, i.e., artificial neural network (ANN) for predicting macromechanical parameters of rock, i.e., UCS based on input cohesive contact shear strength parameters, i.e., cohesion and friction angle, is explored. Results indicate that the ANN model can be quite accurate predictor of uniaxial compressive strength (UCS) and therefore used as a potential tool to aid in estimating target values.

Numerical analysis of the thermally induced damage in remote laser cut carbon fibre reinforced polymers

AbstractRemote laser cutting in carbon fibre reinforce polymers is a promising machining technology which can reduce the main drawback in laser cutting, the heat affected zone. This contribution presents a two‐step modelling approach for the simulation of remote laser cut carbon fibre reinforce polymers, consisting of a damage model for the initially intact material and a model for the thermally damaged material.The static damage behaviour was modelled with the Pinho material model [1], which stands out due to its physically based failure criteria. Initial thermal damages caused by the heat input were modelled with the damage parameters of the Pinho model. The dimensions of the heat affected zone are taken from polished micro sections of the material around the cutting edge.As a numerical example, comparative tensile tests of milled and laser cut open hole specimens were simulated. The advantage of these specimens is the notch effect around the holes in the heat affected zone, which amplifies the differences between the two cutting technologies.

High throughput procedure utilising chlorophyll fluorescence imaging to phenotype dynamic photosynthesis and photoprotection in leaves under controlled gaseous conditions

BackgroundAs yields of major crops such as wheat (T. aestivum) have begun to plateau in recent years, there is growing pressure to efficiently phenotype large populations for traits associated with genetic advancement in yield. Photosynthesis encompasses a range of steady state and dynamic traits that are key targets for raising Radiation Use Efficiency (RUE), biomass production and grain yield in crops. Traditional methodologies to assess the full range of responses of photosynthesis, such a leaf gas exchange, are slow and limited to one leaf (or part of a leaf) per instrument. Due to constraints imposed by time, equipment and plant size, photosynthetic data is often collected at one or two phenological stages and in response to limited environmental conditions.ResultsHere we describe a high throughput procedure utilising chlorophyll fluorescence imaging to phenotype dynamic photosynthesis and photoprotection in excised leaves under controlled gaseous conditions. When measured throughout the day, no significant differences (P > 0.081) were observed between the responses of excised and intact leaves. Using excised leaves, the response of three cultivars of T. aestivum to a user—defined dynamic lighting regime was examined. Cultivar specific differences were observed for maximum PSII efficiency (Fv′/Fm′—P < 0.01) and PSII operating efficiency (Fq′/Fm′—P = 0.04) under both low and high light. In addition, the rate of induction and relaxation of non-photochemical quenching (NPQ) was also cultivar specific. A specialised imaging chamber was designed and built in-house to maintain gaseous conditions around excised leaf sections. The purpose of this is to manipulate electron sinks such as photorespiration. The stability of carbon dioxide (CO2) and oxygen (O2) was monitored inside the chambers and found to be within ± 4.5% and ± 1% of the mean respectively. To test the chamber, T. aestivum ‘Pavon76’ leaf sections were measured under at 20 and 200 mmol mol−1 O2 and ambient [CO2] during a light response curve. The Fv′/Fm′was significantly higher (P < 0.05) under low [O2] for the majority of light intensities while values of NPQ and the proportion of open PSII reaction centers (qP) were significantly lower under > 130 μmol m−2 s−1 photosynthetic photon flux density (PPFD).ConclusionsHere we demonstrate the development of a high-throughput (> 500 samples day−1) method for phenotyping photosynthetic and photo-protective parameters in a dynamic light environment. The technique exploits chlorophyll fluorescence imaging in a specifically designed chamber, enabling controlled gaseous environment around leaf sections. In addition, we have demonstrated that leaf sections do not different from intact plant material even > 3 h after sampling, thus enabling transportation of material of interest from the field to this laboratory based platform. The methodologies described here allow rapid, custom screening of field material for variation in photosynthetic processes.

The experiment of fracture mechanics characteristics of Yellow River Ice

Fracture mechanics provide a reliable approach for describing the transformation of intact ice material into broken and fragmented ice by crack growth. In this work, fracture toughness experiments of natural Yellow River ice were performed with a Brazilian disk method. Physical test results showed that the Brazilian disk test method can be used to determine the fracture toughness and tensile strength of Yellow River ice. With the strain rate increased, the fracture toughness tended to decrease. As the temperature decreased, the fracture toughness tended to increase. In addition, the tensile strength under different temperatures and strain rates was analyzed. The obtained material parameters of Yellow River ice can be used to estimate the fracture strength of Yellow River ice.

Incorporation of micro-cracking and fibre bridging mechanisms in constitutive modelling of fibre reinforced concrete

The formation and propagation of cracks under progressive loading in fibre reinforce concrete (FRC) are significantly influenced by fibre bridging mechanisms. Cracking and fibre bridging, governed by the FRC constituents and their properties, are two coupled and interacting phenomena that significantly affect the ductility and transition from diffuse to localised deformation. Constitutive modelling of FRC is challenging due to the high inhomogeneity and complex transition of deformations stages rooted from the difference in responses of cracked and intact material volumes coupled with cohesive resistance and fibre bridging of a crack. In this paper, a new approach to constitutive modelling of FRC is developed by enriching the constitutive structure to accommodate different responses of the crack, intact material and fibres. The strain discontinuity caused by cracks is accounted for via an enriched strain field which facilitates the introduction of the two interacting mechanisms, cohesive cracking and fibre bridging, in the constitutive model. The transition from diffuse to localised deformation is controlled by the fibre volume content and local deformation, via the density of active cracks. It is demonstrated that the proposed constitutive model is capable of describing the transition from diffuse to localised deformation associated with different macro responses under different loading conditions.

The shear strength of Opalinus Clay shale in the remoulded state

The Opalinus Clay shale formation is considered as a potential host geomaterial for the Swiss deep geological repository for radioactive waste. It presents different facies and it is characterised by a multi-scale heterogeneous composition, by a typical fissile structure with well-defined bedding planes and by anisotropic hydro-mechanical behaviour. This peculiar complexity makes it difficult to assign a unique set of geomechanical parameters to the material. This paper presents an experimental study aimed at characterising the lowest values of the shear strength parameters. In this sense, the shear behaviour was investigated on remoulded samples where the fabric and the diagenetic bonds of the intact material were eliminated. The results of a triaxial test campaign belonging to different facies of Opalinus Clay shale (“shaly” and “sandy” facies) are presented. Furthermore, with the aim to study the mechanical properties of fault zones in Opalinus Clay, the effect of large cumulated shear displacements on shear strength was also investigated. Ring shear tests were performed to determine the residual shear strength. The main geotechnical property which discriminates the different facies of Opalinus Clay is the grain-size distribution; in this sense, well-defined correlations between this intrinsic characteristic and shear strength angles of the remoulded material are presented.

Impact of pulsed electric fields on physical properties of freeze-dried apple tissue

Abstract The aim of this study was to analyze the impact of pulsed electric field (PEF) pre-treatment of apple tissue on kinetics of freeze-drying preceded by vacuum freezing and physical properties of such processed material. PEF intensified freeze-drying kinetics and thus reduced processing time by 57% in comparison to untreated apples slices. Furthermore, the effective water diffusion coefficient increased by 44% as a result of PEF application. Water activity changes during storage of freeze-dried apple tissue were more evident in the case of untreated material albeit initial water activity was higher in the case of electroporated samples. As proved by thermal properties measurements these differences were linked to higher crystallinity of the PEF treated samples (35.5%) in comparison to the untreated material (11.0%). The freeze-dried fruits subjected to PEF pretreatment absorbed more water than the untreated samples while no changes were observed for hygroscopicity and loss of the soluble solids during rehydration. Industrial relevance Obtained results confirm that application of pulsed electric field before freeze-drying makes possible to obtain high quality freeze-dried product preceded by vacuum freezing performed inside freeze-dryer which could reduce freeze-drying time. Since the rehydration capacity of PEF treated material, expressed as ability to absorb water, is better than untreated samples the application of this method could be beneficial when designing instant products. Performed research indicates also that PEF treated freeze-dried apples exhibit better stability expressed by water activity which in turn is related to crystallinity. What more, higher crystallinity of PEF pre-treated freeze-dried samples not only complements the explanation of different physical properties in comparison with intact freeze-dried material but also suggest that sensorial properties may vary.

Multiscale evaluation of potential damage in jetted lateral boreholes

We study the potential formation damage effects due to radial jet drilling on Bentheim sandstone, a homogeneous sandstone that is well established in terms of mechanical properties and widely used as reservoir analogue. We quantify how properties degrade with distance from a jetted hole on μm scale (porosity) and cm scale (compressive, tensile and acoustical properties). Moreover, we perform true-triaxial compression tests on samples with and without a jetted hole. It is concluded that, for this material, jetting has no direct influence on the surrounding area. No significant changes compared to intact material is found, nor can a significant change be detected with respect to distance to a jetted hole. Differences fall within the intra-block variability, and differences between blocks can be well explained by block-to-block variation. Differences in results of the true-triaxial compression tests can be attributed to a different sample geometry, which is corroborated by a numerical simulation. We conclude that the stress field around the jetted hole can therefore be well approximated by the Kirsch equations, modified for compression, and thereby accurately describe the stability of the lateral borehole.

Refined Approaches for Estimating the Strength of Rock Blocks

Micro-discrete fracture networks (μDFNs) have been integrated into grain-based models (GBMs) within the numerical software UDEC to assess rock block strength through a series of unconfined compressive strength (UCS) tests of progressively larger in size numerical specimens. GBMs were generated by utilizing a Voronoi tessellation scheme to capture the crack evolution processes within the intact rock material, and μDFNs were separately created and embedded into the GBMs to simulate the effect of pre-existing defects. Various μDFNs realisations were generated stochastically within the software FracMan to assess the combined impact of defect intensity, persistence, strength and specimen size. The resulting synthetic rock block models were used to assess the “flawed” material strength at block scale through a rigorous sensitivity numerical analysis. The acquired results predict a progressive strength reduction with decreasing intact rock quality and certain trends are captured when rock block strength is expressed as a function of a newly proposed “Defect Intensity× Persistence” factor. This allow us to standardise the data along specific strength reduction envelopes and to propose generic relationships that cover a wide range of defect geometrical combinations, defect strengths and sample sizes. Accordingly, an attempt is undertaken to refine two existing empirical approaches that consider the effect of scale and micro-defects explicitly for predicting the UCS of rock blocks.

The first Adriatic finding of Speleogobius llorisi (Actinopterygii: Gobiiformes: Gobiidae)

In June 2018 two specimens of recently described fish Spelegobius llorisi Kova&#269;i&#263;, Ordines et Schliewen, 2016, were collected at Hvar Island in the central Adriatic Sea at 52 m depth during a SCUBA diving fieldwork, aimed to quantitatively study fishes of rhodolith beds. The findings represent the third known record of this species and the first for the Adriatic Sea. The complete and intact material enabled a study of colouration and morphological details not recorded in the damaged type material from a beam trawl.

Probabilistic analysis of the uplift resistance of buried pipelines in clay

Uplift resistance provided by soil cover is a key aspect in the design of a buried offshore pipeline. The capacity must be sufficient to avoid upheaval buckling but prediction of the uplift resistance is complicated by disturbance of the soil structure during installation. Previous numerical investigations of pipeline uplift capacity in undrained clays have employed simple elasto-plastic models and consider a homogeneous clay. These simplifications neglect critical features of soil-pipeline interaction and may not describe the real mechanical behaviour. In this paper, an advanced kinematic hardening model implemented in a finite element code is used to capture the degradation of structure as a pipeline buried in a natural clay is lifted upwards. The spatial variability of clay structure is represented by a random field and Monte Carlo simulation used to characterise the response. This novel framework shows that clay structure has a significant effect on the failure mechanism and uplift capacity of a buried pipeline. The probabilistic approach accounts for the uncertainty in the condition of the clay backfill and reveals that the spatial distribution of intact and remoulded material can change the mode of failure, emphasising the importance of considering clay disturbance in design.

Detection of unamplified target genes via CRISPR-Cas9 immobilized on a graphene field-effect transistor.

Most methods for the detection of nucleic acids require many reagents and expensive and bulky instrumentation. Here, we report the development and testing of a graphene-based field-effect transistor that uses clustered regularly interspaced short palindromic repeats (CRISPR) technology to enable the digital detection of a target sequence within intact genomic material. Termed CRISPR-Chip, the biosensor uses the gene-targeting capacity of catalytically deactivated CRISPR-associated protein 9 (Cas9) complexed with a specific single-guide RNA and immobilized on the transistor to yield a label-free nucleic-acid-testing device whose output signal can be measured with a simple handheld reader. We used CRISPR-Chip to analyse DNA samples collected from HEK293T cell lines expressing blue fluorescent protein, and clinical samples of DNA with two distinct mutations at exons commonly deleted in individuals with Duchenne muscular dystrophy. In the presence of genomic DNA containing the target gene, CRISPR-Chip generates, within 15 min, with a sensitivity of 1.7 fM and without the need for amplification, a significant enhancement in output signal relative to samples lacking the target sequence. CRISPR-Chip expands the applications of CRISPR-Cas9 technology to the on-chip electrical detection of nucleic acids.