Multiple clinical studies have revealed a link between genomic instability and response to anti-PD-1/PD-L1 therapy in cancer management. A recent study has revealed an important role for the ATR/Chk1 DNA damage checkpoint in regulating PD-L1 expression, raising important clinical and translational questions for therapy selection and study design. A recent report by Sato et al. begins to shed light on the molecular underpinnings of signal transduction from DNA damage to the immune checkpoint, and identifies a key role for the DNA damage sensor kinases ATM, ATR, and Chk1 in this process  was recently approved by the FDA for treatment of patients with advanced MMR-deficient tumours.The association between repair deficiency and improved response to PD-1/PD-L1 blockade has now been demonstrated for several DNA repair pathways. Tumours with mismatch repair (MMR) pathway deficiency exhibit a microsatellite instability (MSI) phenotype, and are characterised by a high mutation burden, an activated immune microenvironment, and increased PD-1/PD-L1 expression on tumour and immune/stromal cells. As such, MSI tumours exhibit high response rates to anti-PD1/PD-L1 therapy,BRCA1, BRCA2, and PALB2) have increased mutation burden, predicted neoantigen load, and PD-1/PD-L1 expression. This raises the possibility that HR-deficient tumours may be sensitive to PD-1/PD-L1 inhibitors; however, their response to these agents appears less robust compared with MMR-deficient or other hypermutated cancers, such as melanomas and non-small cell lung cancers.5 In addition, tumours with mutations in the exonuclease (proofreading) domain of polymerase epsilon (POLE) or polymerase delta (POLD1) have among the highest mutation burdens identified to date, and POLE/POLD1-mutated tumours exhibit high levels of immune activation and PD-1/PD-L1 expression, and are sensitive to immune checkpoint blockade.7Associations between alterations in other DNA repair pathways and tumour immune features have also been reported. Tumours with modifications in the homologous recombination (HR) pathway ] resulted in the largest increase in radiation-induced PD-L1 expression. Again, they observed that pharmacologic inhibition of ATM, ATR, or Chk1 was sufficient to abrogate PD-L1 upregulation. Interestingly, PD-L1 upregulation in Ku80-depleted cells could also be abrogated by depletion of either EXO1 (exonuclease) or BLM (helicase); two genes that are required for DSB processing and end resection. Similarly, in their screen, loss of BRCA1 did not show a significant increase in ionizing radiation-induced PD-L1 expression, compared with control cells. This highlights the importance of Chk1 activation via end resection as a key process in DSB-induced PD-L1 expression. Although both BRCA1 and BRCA2 are required for HR, they have distinct roles in this process; BRCA1 promotes DNA end resection by relieving the barrier posed by 53BP1 in HR, a process where BRCA2 is not involved. Therefore, according to their model, DNA end resection is impaired in BRCA1-defective cells, meaning ATR/Chk1 signalling is not effectively activated and PD-L1 activation does not occur. This is in stark contrast to BRCA2-defective cells where DNA end resection is not impaired as BRCA2 is not involved in this process. Finally, the authors showed that damage-induced ATM/ATR/Chk1-mediated PD-L1 upregulation is dependent on IRF1 signalling through phosphorylated STAT1/3.Despite these robust clinical associations, the mechanisms linking DNA damage to increased PD-L1 expression and immunotherapy response are incompletely understood. To functionally characterise the impact of DNA damage on PD-L1 expression, Sato et al. subjected several DNA repair-proficient cancer cell lines to ionizing radiation, double strand break (DSB)-inducing drugs such as etoposide, or a poly(ADP)-ribose polymerase inhibitor (PARPi). Treatment resulted in a time-dependent increase in PD-L1 expression at both the transcriptional and protein levels. To investigate the role of the DNA damage response in this process, they added specific inhibitors of ATM, ATR, or Chk1, and demonstrated that inhibition of any of these DNA damage checkpoint kinases could suppress PD-L1 upregulation. Next, the authors screened an siRNA library targeting DNA repair genes; they found that depletion of The work by Sato et al. provides important mechanistic insights linking DNA damage with immune activation, by identifying a critical role for the DNA damage checkpoint in regulating PD-L1 expression. Although these data have provided a window into one aspect of the signalling network that links genomic instability with immune signalling, the molecular details of much of the network remain to be elucidated. Particularly, the events downstream of ATR activation, including transcriptional changes and/or direct activation (or repression) of other signalling factors, may uncover additional mechanisms through which the immune response can be modulated by DNA damage.BRCA2-/- cells) are unclear. In addition, many of these experiments will ultimately have to be conducted in immune-competent model systems in order to fully understand the breadth of the immune response to DNA damage.Temporal aspects of DNA damage (or repair deficiency) may also impact the immune response: in the experiments by Sato et al., changes in PD-L1 levels were measured following acute damage exposure or siRNA-mediated depletion of a repair factor, and the observed PD-L1 increase did not persist beyond 14 days. Therefore, the dynamics of PD-L1 expression following chronic damage exposure or in a constitutively DNA repair-deficient background , radiation therapy, or DSB-inducing chemotherapy agents.1 The data by Sato et al. raise the possibility that, at least in the setting of exposure to DSB-inducing agents, DNA damage checkpoint inhibitors  may decrease tumour response to immune checkpoint inhibition by suppressing PD-L1 expression, thereby arguing against a triplet of DSB-inducing therapy/ATR-Chk1 blockade/PD-1-PD-L1 blockade. Nonetheless, the optimal sequence of conventional or targeted agents with anti-PD1/PD-L1 therapy may be highly context-dependent, and this work highlights the important role for pre-clinical studies in identifying potential mechanisms of synergy or antagonism.This work also has several important clinical implications. The optimal timing and combinations of DNA damaging agents with immunotherapy are not known. Furthermore, both immune stimulating and suppressive effects of combined regimens have been noted.BRCA1-mutated tumours, resistance to PARPi may occur due to a rescue of DNA end resection ability via loss of 53BP1, REV7, or Ku70/80, which increase HR capacity.11 In this setting, re-establishing end resection may promote PD-L1 upregulation via EXO1/BLM (and ATM/ ATR/ Chk1) activity, and thus sensitise tumours to PD-1/PD-L1 blockade with or without DSB-inducing agents. In BRCA2-mutated tumours, resistance to PARPi may occur via protection of the replication fork,13 which is dependent on ATR activity.14 This ATR activity may lead to upregulation of PD-L1 and promote response to PD-1/PD-L1 blockade. In both scenarios of PARPi resistance in BRCA1/2-mutated tumours, the study by Sato et al. suggests that PD-1/PD-L1 inhibitors may be a useful therapeutic strategy (with or without concurrent DNA damaging agents) for tumours that have progressed on PARPi.Another clinical implication of this work is the selection of anticancer therapy after PARPi progression. PARPi are synthetically lethal with HR deficiency and are now FDA approved for clinical use in ovarian cancer, while also being evaluated in several other HR-deficient tumour settings. However, a substantial fraction of patients eventually develop resistance to PARP inhibition and several mechanisms of PARPi resistance are now emerging. In The work by Sato et al. highlights an important role for the DNA damage ATR/Chk1 checkpoint in regulating PD-L1 expression, thus linking DNA DSB signalling with regulation of the immune response. These observations have important clinical implications for therapy selection, particularly following progression on PARPi and other DNA damaging agents. They also have translational implications for the design of appropriate correlative studies in ongoing and future clinical trials of DNA damaging agents in combination with immunotherapy.


We propose a broadband terahertz absorber consisting of nonstructured graphene loaded with arrays of elliptic dielectric cylinders. The relative bandwidth for the absorption above 90% reaches about 65%. The working mechanism of broad bandwidth mainly comes from two aspects. One is that the nonstructured graphene loaded with elliptic dielectric cylinders provides multiple discrete graphene plasmon resonances with large relative frequency interval. The other is that, for each discrete resonance, there exists a set of continuous plasmon resonances because the width of the dielectric structure varies continuously and gradiently. The broadband terahertz absorber we demonstrate here, based on geometrically gradient dielectric structures and nonstructured graphene, avoids the graphene processing, which shows great potential applications in related devices. Many applications of THz devices have been proposed so far, such as communicating, spectroscopy, sensing and imaging4. Among these attempts, the absorber plays an important role in many devices working in THz range, including detectors5, sensors6, modulators7, thermal emitters8, camouflage devices9, etc. Traditionally, metallic metamaterials are the key components of many THz absorbers, structures of various geometries have been designed to realize different absorption functions such as dual-band11, multi-band13 and broadband16. Recently, graphene, a single-layered carbon atom arranged in a honeycomb lattice, has become one of the most promising materials for designing THz absorbers due to its tunability, broadband response and high carrier mobility17. Various types of graphene-based absorbers have been proposed to achieve near-unity absorption. Many of these absorbers are based on periodically structured graphene like disks18, microrings19, fishnets20, ribbons with gradient width21 and multilayered ribbons with asymmetric voids22. Nevertheless, edge effects of structured graphene are evident disadvantages of these structures23. To avoid this problem, absorbers using nonstructured graphene are put forward, such as metal-dielectric-graphene sandwich structure24, graphene loaded with periodical arrays of dielectric bricks25, multilayer graphene sheets on quartz substrates26, graphene with metasurface comprising of plasmonic structures27, and multilayer graphene with uneven dielectric slab structure28.Terahertz (THz) devices have attracted increasing attention due to the unique properties of the THz region (0.1–10 THz), which share similarities with both the microwave and the far infrared rangesIn this letter, we propose an alternative broadband mechanism of multi-band continuous plasmon resonances, and demonstrate that the relative bandwidth of over 90% absorption can reach about 65% in the THz range by using nonstructured graphene loaded with geometrically gradient dielectric structures. Here, we expound the physical mechanism firstly. Then, we present numerical simulations and results. Finally, we investigate the effects of some relative parameters.W along x-axis, the periodic interval L along y-axis, the semi-minor axis R1, the semi-major axis R2 and the thickness H2 of the elliptic dielectric cylinder, the Fermi level EF of graphene, the thickness H1 of the dielectric substrate and the thickness Hm of the metallic film. The material of the dielectric substrate and the elliptic dielectric cylinder are dielectric 1 and dielectric 2, respectively. In this structure, the metallic film can reflect waves and the nonstructured graphene loaded with periodic arrays of geometrically gradient dielectric can also provide array local resonant reflection (ALRR)29, which form two mirrors of an asymmetric Fabry-Perot (FP) cavity30. The transmission of waves can be blocked as long as the metallic mirror is thick enough compared with the typical skin depth at THz frequencies. The reflection of waves can be eliminated if the critical coupling condition is satisfied27 or the impedance of the absorber is matched to that of the free space31. When both the transmission and reflection channels are suppressed, the near-unity absorption can be realized. Since the near-unity absorption is decided by the critical coupling condition or impedance matching condition that is closely related with the graphene plasmon resonances (GPRs), we can broaden the absorption band through broadening GPRs. However, generally, the GPRs are narrowband. So, in order to achieve broadband absorption, the key problem is to find a method to broaden the GPRs. In this configuration, the GPRs are related to the formation of standing-wave patterns of graphene plasmons with the wave vector33.e is the charge of electron, ħ is the reduced Planck’s constant, ω is the angular frequency of graphene plasmons, EF is the Fermi level of graphene, τ is the carrier relaxation time in graphene, ε0 is permittivity of vacuum, ε01 is the relative permittivity of the dielectric substrate, and ε02 is the relative permittivity of dielectric cylinders above the graphene film. When the structures are illuminated by a plane wave with the electric field linearly polarized along the x direction, the excited graphene plasmons propagate along the x direction and experience multiple reflections from the interfaces between the dielectric cylinders and air gap. In this structure, based on the thin element approximation in scalar Fourier optics, the elliptic dielectric cylinders can be divided into infinite infinitesimals with the profile of isosceles trapezoids along the y-axis, as shown in Figure Lp(y), namely the FP cavity lengths, between two mirrors at a specific position y is approximated askGP) is the real part of wave vector kGP, N is a positive integer determining the order of a resonance mode, ϕ is the phase of the reflection coefficient for graphene plasmon reflection at two mirrors. In general, ϕ is non-zero that depends on the structural, material parameters and position y. The physical explanation for this non-zero f is related to extension of the plasmon field beyond the interfaces of the dielectric cylinders and air gap.The proposed structure of the broadband THz absorber is presented in Fig. N can be set as different positive integers. We can also see that, for each discrete resonance, there exists a set of corresponding continuous plasmon resonances in the structure because y varies continuously in the elliptic dielectric cylinders. Under certain conditions, the frequency region of the continuous resonances can be larger than the corresponding frequency interval between two discrete neighbor resonances. So, the overlapping resonant frequencies can spread over a wide range, which is referred to as multi-band continuous GPRs.From eqs –3), by , by 3), β between two mirrors of a FP cavity is a key factor determining the performance of resonance, and an angle of zero or close to zero is a condition for obtaining strong resonances. In our proposed structure, the angle β between two mirrors at a specific position y can be approximately given byAs we know, the angle R1 and R2, the angle β decreases with a decrease of position y, which shows the FP-type resonances are most likely to occur at smaller position y and can always occur at position y = 0 particularly. For a fixed R1 and position y, the angle β becomes smaller and tends to zero as R2 increases. This implies that it is reasonable to use the elliptic dielectric cylinders rather than usual dielectric cylinders to act as the geometrically gradient dielectric to obtain multi-band continuous GPRs. Since the near-unity absorption is closely related with the GPRs, the achieved multi-band continuous GPRs imply that the broadband near-perfect absorption may be obtained under certain conditions.From equation , we can εr1 = 4 (dielectric 1) and εr2 = 12 (dielectric 2) are selected to show the physical mechanisms of broadband absorption. In the THz range, εr1 = 4 and εr2 = 12 can be approximatively provided by silicon dioxide and silicon, respectively34. The metallic material in this structure is gold, which is described by Drude model. The relative permittivity of gold is expressed by ε∞, ωP and γ are 1.0, 1.38 × 1016 rad ⋅ s−1, and 1.23 × 1013 s−1, respectively29. Graphene is modeled as an anisotropic layer with thickness hg = 1 nm. The relative permittivity of graphene comprises the out-of-plane component εout = 2.5 and the in-plane component εin = 2.5 + iσ(ω)/(ωε0hg), where the graphene optical conductivity σ(ω) is derived using the random-phase approximation (RPA)36. σ(ω) is calculated by σ(ω) = σinter(ω) + σintra(ω), where σinter(ω) and σintra(ω) are the interband and intraband contributions, respectively. In the THz range, the interband part σinter(ω) is negligible compared with the intraband part σintra(ω)37, hence we can use σintra(ω) to approximate σ(ω) here, which is described asω is the frequency of the incident wave, kB is the Boltzmann constant, e is the charge of an electron, T = 300 K is the temperature, & planck; is the reduced Planck’s constant, τ is the carrier relaxation lifetime, and EF is the Fermi level. Here, relaxation lifetime is expressed by 27, where the mobility μ is 10000 cm2/(V ⋅ s) and the Fermi velocity vF is 106 m/s. In the simulation, the open boundary condition is adopted in z direction and the unit cell boundary condition is adopted in x and y directions. The absorption is calculated from the obtained S parameters using A = 1 − R − T, where R is the reflection and T is the transmission.To verify the theoretical prediction, we next conduct full-wave numerical simulations employing frequency domain solver in CST Microwave Studio. In the simulation, EF = 0.5 eV, H1 = 15 μm, H2 = 11 μm, Hm = 2 μm, R1 = 10 μm, R2 = 36 μm, L = 80 μm, W = 24 μm. Figure First, we consider the case where R2, incident angle θ and Fermi level EF. It is necessary to stress that in each group of simulations, all the other parameters and conditions are kept the same as the above simulations.In order to investigate the characteristics of the broadband THz absorber, we sweep a few parameters, including semi-major axis R2 from 1 to 40 μm while fixing R1 = 10 μm. Figure R2/R1 under normal incident wave with the electric field parallel to x-axis. It can be observed that the absorption is very sensitive to ellipticity R2/R1. When the values of R2/R1 are very large (e.g. R2/R1 > 3), the characteristics of broadband absorption maintain well. On the other hand, when R2/R1 is relatively small (e.g. R2/R1 < 1.5), the broadband absorption tends to be torn. This is in agreement with that, decreasing the ellipticity R2/R1 results in an increase of the angle β between two mirrors of FP-type GPRs at specific positions in the elliptic dielectric cylinders, which eventually makes the corresponding resonance strength weaker and even disappearing. As a result, the corresponding absorption becomes small and the broadband absorption is torn. In order to further confirm the broadband mechanism, the typical distributions of the z component of the electric field at the distance 50 nm above the interface between the graphene and the elliptic dielectric cylinders for R2/R1 = 1 and R2/R1 = 0.4 are shown in Fig. R2/R1 = 3.6. Comparing the typical distributions of electric field in three different R2/R1 ratios, we can visually find that the larger R2/R1 can really provide more GPRs. For example, at the case of R2/R1 = 0.4, the FP-type GPRs only occur at the position y = 0. However, for R2/R1 = 3.6, the FP-type GPRs can occur at a wide region of position y. These field distributions are consistent with the prediction.We first investigate the effect of ellipticity of elliptic dielectric cylinders by varying θ from 0° to 89.9° while maintaining the incident plane wave in the x-z plane and the magnetic field parallel to y-axis to examine the incident angular sensitivity. The incident angle θ is defined as the angle between the incident plane wave and the positive z-direction. Figure θ. It can be seen that the dependence of the absorption on the incident angle is relatively weak when the incident angle varies between 0° and 60°. The reason for this is that the near-unity absorption of this type of absorber is closely related to the GPRs. However, the excitation of the GPRs is insensitive to the incident angle38. As the incident angle increases beyond 60°, the absorption decreases rapidly. This can be understood through the following qualitative analysis. The near-unity absorption is also closely related to destructive interference. But, in the case of large angle incidence, the reflection amplitudes from two mirrors of the asymmetric FP cavity formed by the metallic film and the nonstructured graphene loaded with periodic arrays of geometrically gradient dielectric have a large mismatch, and the destructive interference amplitude-matching condition is destroyed. It is noticeable that there is an abnormal absorption phenomenon near the incident angle 85°. The reason of this phenomenon is: for the structure consisting of a single layer of lossy two-dimensional (2D) material separated from a mirror by a dielectric spacer layer, there always exists an angle of incidence, at which the external radiation decay equals to internal absorption decay and thus complete absorption of the 2D material can be achieved39. For our absorber, at 3.6 THz, the above condition of complete absorption is satisfied when the incident angle is near 85°.Next, we vary the incident angle EF on the absorption in Fig. EF increases, which is because the plasmon resonance frequencies of the doped graphene increase with increased Fermi level40. It can also be seen that the absorption decreases rapidly when EF varies from 0.2 to 0 eV, which results from the fact that the metallic character of graphene decreases when EF is relatively small. However, there is an interesting and abnormal phenomenon that a narrowband absorption still exists near the upper absorption band even as EF approaches 0, where GPRs do not exist. To explain this phenomenon, we extract the absorption for EF = 0.0 eV from Fig. Then, we show the effect of Fermi level In summary, we propose a broadband THz absorber based on multi-band continuous plasmon resonances sustained by geometrically gradient dielectric loaded with graphene. The simulation results show that the relative bandwidth for the absorption above 90% can reach about 65%. The broad bandwidth mainly originates from two key aspects. One is that the nonstructured graphene loaded with geometrically gradient dielectric can provide multiple discrete graphene plasmon resonances with large relative frequency interval. The other is that, for each discrete resonance, there exists a set of continuous plasmon resonances because the width of dielectric structure varies continuously and gradiently. Besides, FP resonances of the dielectrics also contribute to the absorption. Based on nonstructured graphene, the absorber avoids processing graphene and destroying the unique properties of graphene. The approach of combining geometrically gradient dielectric structures and nonstructured doped graphene facilitates the application of the broadband THz absorbers and related devices.


Porous graphene has been proposed as superior material for highly selective membranes for gas separation. Here we design some models of porous graphene with different sizes and shape as well as employ double layers porous graphene for efficient CO2/H2 separation. The selectivity and permeability of gas molecules through various nanopores were investigated by using the reactive molecular dynamics simulation which considers the bond forming/breaking mechanism for all atoms. Furthermore, it uses a geometry-dependent charge calculation scheme that accounts appropriately for polarization effect which can play an important role in interacting systems. It was found that H-modified porous graphene membrane with pore diameter (short side) of about 3.75 Å has excellent selectivity for CO2/H2 separation. The mechanism of gas penetration through the sub-nanometer pore was presented for the first time. The accuracy of MD simulation results validated by valuable DFT method. The present findings show that reactive MD simulation can propose an economical means of separating gases mixture.The separation of gases molecules with similar diameter and shape is an important area of research. For example, the major challenge to set up sweeping carbon dioxide capture and storage (CCS) in power plants is the energy requisite to separate the CO Because of increasing demand for purified hydrogen, extensive research studies have been devoted to the advancement of the technologies for generation and purification of hydrogen with higher efficiency and lower production cost6.Due to its abundance, easy synthesis, and nonpolluting properties, hydrogen has been considered to be the most promising energy source, nowadays. Hydrogen energy technologies can play a special role in solving problems related to the environment sectors2 streams with different compositions, which require corresponding CO2 separation processes. CO2 a main greenhouse gas (GHG), is discharged into the atmosphere via different processes: namely the fossil fuel combustion 7. Accounting for about 60% of the heat trapped in the atmosphere, CO2, potentially, has the greatest inclusion in climate change, among all GHGs. The increase in air temperature along with changes in precipitation patterns, has led to an increase in the number and strength of natural disasters; like floods, droughts, hurricanes, extensive melting of ice and snow, and an increase in medium sea levels8. These affect directly and indirectly on human life and ecological systems on the planet earth. Increasing the general consciousness on the subject has urged researchers and policy makers, to study climate change and find a solution to lessen this threat.The universal carbon dioxide emissions, on the other hand, are expected to increase as long as fossil fuels continue to be the main energy source for mankind. Numerous industrial processes emit CO2 capture and storage (CCS) have the most importance. In spite of researches about renewable energy, the fossil fuels are the main energy for mankind and remains as predicted, the main energy for at least the next two decades. So, CCS is the best way to lessen climate change9. CO2 capture is currently a topical issue in environmental protection and sustainable development10 and has been intensively researched and developed to survive with climate change, by reducing atmospheric CO2 concentration12. CO2 can be obtained from numerous sources including fossil fuel power plants, refineries, oil and gas production sites, iron and steel factories, cement and other chemical plants. CO2 capture can be classified into four main techniques: 1- pre-combustion 2- post-combustion 3- oxy-fuel and 4-electrochemical separation16. Pre-combustion capture uses new gasification method to create combustible gas and then capture the CO2 before burning for power. In pre-combustion capture, the CO2 concentration is high, which lead to a higher driving force for the CO2 separation and potential for cost savings in the absence of compression necessity. For pre-combustion capture, the carbonaceous fuel is first converted to a form amenable to CO2 capture.There are a number of ways to alleviate the GHGs emission to the atmosphere that among them replacing common fossil fuel based energies with more environmentally compassionate sources (renewable energy), and the CO2 and CO2 with a trace amount of H2O and H2S gases. The high concentration of CO2 (20–40%) in the fuel gases needs to be eliminated before the combustion. The syngas gas is a mixture of CO, H2, H2O, CO2, and other components. There are some impurities in the syngas, which need to be eliminated and then through a two-step water gas shift, carbon monoxide would convert to carbon dioxide17. The produced mixture is mainly composed of CO2 and H2. Then the mixture goes through a CO2 capture system, to create a nearly-pure hydrogen stream. This hydrogen flow goes into a combined cycle power plant to generate electricity.The fuel gas called synthetic gas (syngas) is composed of mainly H2 capture: a) adsorption, b) absorption, c) cryogenic distillation, d) membranes, e) gas hydrates, and f) chemical looping18. Except for cryogenic separation, the rest of these methods require some material as carriers. Currently, absorption, via solvent scrubbing, is a deep-rooted CO2 separation approach, which has been implemented broadly in chemical and petroleum industries. Cryogenic distillation is based on the principales of separation based on cooling and condensation. It has been used in liquid separations for a long time. This technique is hypothetically very useful for CO2 separation; but the substantial energy requirement makes it less desirable for most applications. Membranes separate gases based on the contrasts in physical and chemical interactions between different gases and the membrane materials. This would allow some components to pass preferentially through the membranes based on size (kinetic) and affinity (thermodynamics)19. Membrane technology can be a powerful tool for CO2 capture process by decreasing equipment size and lowering energy requirement.There are different separation techniques in detail, for CO8. Traditional systems for gas separation in industry consume a very high energy cost4 and lead to environmental problems. The use of membrane system20 as an efficient separation for the gas mixtures without any phase change clearly lessens the energy cost compared to traditional systems. The main advantages that membrane technology presents over other gas separation methods like pressure swing adsorption and cryogenic distillation are its low power usage and costs, simplicity in operation and its compactness and portability1. The good separation membrane must have controllable pore size21, stable structure, and efficient permeability. Numerous studies have been devoted to evaluate potential applications of graphene for suitable gas separation26. Recent theoretical work by Jiang et al. has proposed porous graphene27 as a promising material for highly selective membranes. Graphene is an extended honeycomb network of sp2-hybridized carbon atoms and the first example of a close-packed two-dimensional (2D) crystalline material isolated in nature and has emerged to become an exciting new nanomaterial of carbon with many novel properties29. So, graphene, the single-atom-thick planar membrane29 and the strongest structure30 that exists so far in the earth, becomes interesting for its potential applications to be a good membrane. The use of graphene as gas storage has also been an active area of research, due to their extended π-conjugation and high surface area32.Generally, for industrial gas separation there are three major processes: pressure swing adsorption (PSA), cryogenic distillation and membrane gas separationet al.33 found that gas mixtures can effectively been separated with porous graphene via the size exclusion using sub-nanometer pore rims. Also, the chemical functionalization of graphene pore rims is effective on gas permeability and selectivity. Xue et al.34 demonstrated that porous graphene membrane with N-passive pores has a strong electrostatic force for CO2 compared with N2 when permeating through the pores. In 2016 Wang et al. used both molecular dynamic simulations and DFT calculations to investigate the gas separation performance of H-passive nanoporous graphene membrane. They considered a mixture of CO2/N2 gas molecules and three different pore sizes for the H-passive membrane. They found that H-pore-13 (among H-pore-10 and H-pore-16) has a higher interaction energy with CO2 in compared with N2. Also the barrier energy of CO2 (0.19 eV) is much more than that of N2 (0.05 eV) so N2 can permeate through H-pore-13, whereas CO2 cannot35.The gas separation performance of porous graphene membrane can be investigated by creating the sizes and shapes of their pores and by the chemical functionalization of graphene. For example, Liue 2/H2 gas mixture. We employ reactive molecular dynamics simulation  and DFT calculations in order to gain insights into the penetration and interaction properties of passing molecules through the pore. Our results demonstrate that permeability and selectivity of considered pores are highly sensitive to the functional group atoms as well as the pore size. It was found that the H-passive pore graphene membrane with diameter of 3.75 Å exhibits a high selectivity for CO2/H2 separation. The mechanism of penetrated molecules across the considered pore has also been investigated and validated by using the dispersion corrected DFT calculations. It is expected that this study could provide useful information for CO2 gas capture/separation from H2 as well as environment protection.The current study entails the permeability and selectivity of pore graphene with different functional atoms (C and H) and different pore size for CO2/CO2 gas separation using both quantum mechanical calculations and molecular dynamics (MD) simulations. DFT calculations were employed to optimize the structure of graphene and explore the energy barriers and potential energy surface of H2/CO2 molecules passing through sub-nanometer pores created in a graphene sheet. DFT calculations were performed using the ab initio package SIESTA, which is based on the localized basis set and the method of pseudo potentials33. In this software the electronic wave functions are constructed by linear combination of locally-confined atomic orbitals (LCAO’s)36. Norm-conserving pseudo potentials of Troullier–Martins37 with the valence electron configurations of all considered atoms were implemented. Dispersion interactions should be important for the systems under study and therefore we utilize the DFT-D2 method in order to consider the respective interactions. The DFT-D approach has been extensively tested on numerous systems38 including the physisorption of small molecules to graphene sheets40 and the adsorption of H2 within metal-organic framework materials41. The optimization procedure has been carried out with SCF tolerance of 10−5 eV and residue force on each atom was set to 0.02 eV/Å. The Monkhorst-Pack approach was used to represent the Brillouin zone by k-points of 5 × 5 × 1 meshes for the graphene sheets. We utilized a vacuum thickness of 25 Å along the z direction of the graphene layers. All the atomic coordinates including hydrogen and carbon atoms of the pore membranes were fully optimized.The performance of an idealized graphene membrane was evaluated for HEint, and the corresponding barrier energies, Ebarr are calculated from following equations:EGraph and EM correspond to total energies of the relaxed graphene and isolated H2/CO2 molecule, respectively. EM/Graph is the total energy of the optimized complex. Eint.max and Eint.min are defined as maximum and minimum values of the interaction energies.The values for the interaction energies, 43 calculations were performed by using the modern electronic structure package, ORCA44. We used the def2-TZVP  basis set45 for all atoms. The Grimme approach using atom pair-wise additive schemes46, so-called DFT-D3 method, was utilized to consider the dispersion corrections for the long range non-bonding van der Waals (vdW) interactions. The DFT-D3 method has been shown to be well performed with Becke-type exchange such as BLYP or B3LYP functionals47.In some cases, to verify geometry parameters as well as adsorption properties of interacting entities the hybrid density functional B3LYP40. GULP supports geometry optimization and MD simulations of molecules, clusters, and 2D-/3D-systems using a wide range of potential models that span both the inorganic and organic fields. The potentials include shell model, embedded atoms , and bond order/reactive force fields. The range of potential models means that structures as diverse as zeolites and metallic films on semiconductor substrates can be modeled.Molecular mechanics (MM) calculations with model potentials were used to calculate gas passing barriers and compared with the DFT calculations to assess the accuracy of the classical potentials. All MM calculations were carried out with General Utility Lattice Program (GULP) 4.0 to simulate separation of gas mixturesNo discontinuities in energy or forces, even during reactions.sp2 and sp3 atoms for carbon; the method determines the atoms hybridization from its chemical environment.Each element is described by just one force field atom type. The ReaxFF metal oxide oxygen is described by the same parameters as the ReaxFF oxygen in organic molecules. ReaxFF does not have separate 42.No pre-definition of reactive sites is necessary: given the right temperature and chemical environment reactions will happen automaticallyWe used a superior reactive force field so-called as ReaxFF. At the center of the ReaxFF potential lies a bond order/bond energy relationship. Bond orders were obtained from interatomic distances and continually updated at every iteration allowing for connectivity changes. These bond orders are incorporated in all valence terms  ensuring that energies and forces associated with these terms go to zero upon dissociation. Furthermore, ReaxFF describes non-bonded interactions between all atoms, irrespective of connectivity. Excessive short-range repulsive/attractive non-bonded interactions are circumvented by inclusion of a shielding term in the vdW and coulomb interactions. ReaxFF aims to provide a transferable potential, applicable to a wide range of chemical environments. To ensure its transferability, the following general guidelines were adopted:36, nitramines37, ceramics40, (Si/SiO2), metals and metal oxides38, metal/hydrocarbon interactions37, and metal hydrides40 demonstrating that ReaxFF has the versatility required to capture the complexity of the mixed metal catalyst system. Meanwhile, the ReaxFF potentials are demonstrated to be about two order of magnitude more expensive than conventional force fields such as CHAARMS and AMBER, but are remarkably faster  than ab initio calculation methods.ReaxFF studies have been reported for a wide range of materials, including hydrocarbons2 and 50 H2 molecules in the simulation system with the dimension of 18 Å × 22 Å × 120 Å and periodic boundary conditions (PBC) were applied in all three dimensions. The gas box involved H2 and CO2 gas molecules and two pore graphene sheets were located in both sides of the gas mixture to enhance the efficiency of gas permeability of nanoporous membranes. The porous graphene membranes were located in the middle of the simulation box. It should be noted that there is vacuum space besides two sides of the graphene membranes. The molecules of each gases have been randomly positioned within the simulation box between the nanoporous graphene membrane. The length and width of modeled simulation box match to the length of each side of the nanopore graphene membrane (~2 nm). To avoid displacement of the graphene layers due to the pressure exerted by the gas molecules, the position of two carbon atoms in the sheet have been fixed during the simulation procedure. The selected atoms are located as far away as possible from the pores to simulate realistic behavior of atoms in the neighborhood of the rim.In the present work, there are 50 COns MD simulation, and data were collected at each 0.05 ps. The total energy and the temperature of systems were found to be stable over 20 ps.We performed a constant-volume/constant-temperature dynamics (NVT) simulation at 300 K controlled by the Andersen thermostat method, with a fix time step of 0.5 fs. The vdW interactions were applied with a cut off distance of 12 Å. The Ewald sum scheme was employed to describe the electrostatic interactions. Every modeled system was simulated for a 4 2, CO2 and graphene systems by molecular mechanics based ReaxFF potential and compared the results with the DFT-B3LYP methods. For this aim, a graphene flake consisting of 32 C and 14 H atoms as a suitable model for graphene was considered. Full structural optimization of all molecular systems was performed and the results indicate that there is good agreement between the two calculation results. The structural parameters obtained with considered methods are listed in Table 50,  is the ratio of the number of permeation events of the two types of gas molecules. It can be inferred that if the permeation ratio equal to one, there is no selectivity and the higher ratio means higher selectivity. Pore size was specified as the mean of the shortest and largest inner distances in the pore and the pore area was calculated according to the area of benzene rings drawn out24 . After 4 ns MD simulation the number of passing molecules across the membrane is estimated and listed in Table 2 molecules for CO2/H2 mixture. We next investigate the influence of pore size on the separation process. Different pore sizes of porous graphene saturated with hydrogen are considered  and O atoms of CO2 molecules which causes the CO2 to migrate toward the pore while these positive sites repel the H2 molecules from the pore edge.We first consider the pore 10CH membrane on CO2 passing through pore 16CH porous graphene. According to the MD simulation observation, we can divide the permeation process into three steps. At the first, the molecule moves close to the membrane pore. Then it shifts and moves back and forward several times for some picoseconds, and then goes to the other side of the membrane. During the simulation time, both CO2 and H2 molecule approach the graphene surface. After that one CO2 molecule moves back and forth around the pore and finally overcomes the barrier energy and permeates vertically through the pore at 43 ps of simulation time. The snapshot of passing progress of a CO2 molecule through the pore 16CH is represented in Fig. 2 molecules pass through the pore while after 10 ns of simulation time there is still no H2 transition which shows really high selectivity of 16 H pore for CO2/H2 separation. We have also addressed the changes in the pore conformation when the gas molecules passed through the membrane. To this end, the distances between two vertical/horizontal hydrogen atoms as well as the angle between three C atoms around the pore during the simulation time have been calculated  and 10 ps (CO2 molecule is trapped in the pore). Figure 2 was enhanced when it locates between the H atoms of the pore. Indeed, a significant charge has been transferred from the H atoms of the rim to the O atoms of the trapped CO2 molecule in the pore during the penetration process.To gain further insight into the interaction nature of system under consideration we dedicate the charge transfer between the passing molecule through the pore and the adjacent atoms of the rim. Thanks to the ReaxFF potential employed in the present work, it is now possible to evaluate the amount of transferred charges during the penetration process. The QEq scheme was employed for charge analysis between passing CO2/CO2 gas molecule passing through the center of the pore; i.e., parallel and perpendicular orientations for molecular axis of approaching molecule toward the center of the pore. We considered 16CH pore as the optimal pore for the penetration mechanism. Figure 2 molecule approaching the graphene pore. The calculated interaction energies are given in Table first-principles calculation results show that CO2 molecule prefers to pass through the pore 16CH with its molecular axis perpendicular to the graphene surface. The calculated Ebarr for CO2 molecule passing through the pore with perpendicular orientation was 0.17 eV while in the parallel one was about 0.19 eV  of 3.75 Å (16CH pore) performs high selectivity and desirable permeability for CO2/H2 separation while smaller and larger pores demonstrated less permeability and selectivity, respectively. The 16CH pore was found to be optimal porous membrane among the considered pores and then 13CH pore (short side of about 3.5 Å) with rather the same selectivity but less permeability. The reactive MD simulations demonstrated that CO2 molecules were trapped into the pore and have a long delay time during the penetration due to the attraction between O atoms of the CO2 and H atoms of the pore. Charge analysis by ReaxFF based QEq scheme which is validated by DFT-NBO level of theory indicated that significant charges were transferred from the H atoms of the pore to the O atoms in the trapped CO2 molecule during the penetration process. This reveals rather strong interaction between passing CO2 molecules and pore membrane.In summary, by drilling the graphene lattice and functionalizing the pore rim as well as considering double layers pore strategy, we significantly enhanced the selectivity and permeability of nano-porous membranes for COfirst-principles calculations it was found that CO2 prefer to pass through the 16CH pore with its molecular axis perpendicular to the graphene pore. However, the difference between calculated barrier energies of CO2 and H2 is high (less barrier energy for H2 molecule) while the results showed high permeability to CO2 molecules and no H2 molecules allowed to permit. This observation attributed to higher interaction energies and so higher attraction between CO2 molecules and H atoms of the pore which cause the CO2 molecules to be adsorbed to the pore rim. Finally, reinforce pressure of other gases molecules as driving force push these trapped molecules to easily penetrate through the pore.The mechanism of gas separation was investigated within DFT-D calculations. Using the 2 separation have numerous applications such as industrial applications, CCS (CO2 capture and storage), and environment protection. Future works are dedicated on developing novel hetero atoms nanosheets such as h-BN and ZnO mono-layers as well as functionalized graphene and applying them in gas separation.Therefore, modified pores with 3.75 Å of size (short side) accompanied with double layer strategy provide a promising approach for constructing new materials with innovative separation properties. Researches on CO


LINE-1 (L1) retrotransposons are a notable endogenous source of mutagenesis in mammals. Notably, cancer cells can support unusual L1 retrotransposition and L1-associated sequence rearrangement mechanisms following DNA damage. Recent reports suggest that L1 is mobile in epithelial tumours and neural cells but, paradoxically, not in brain cancers.Alu-Alu recombination event adjacent to an L1. These mutations included PCR validated intronic events in MeCP2 and EGFR. Despite sequencing L1 integration sites at up to 250× depth by RC-seq, we found no tumour-specific, endonuclease-dependent L1 insertions. Whole genome sequencing analysis of the tumours carrying the MeCP2 and EGFR L1 mutations also revealed no endonuclease-dependent L1 insertions. In a complementary in vitro assay, wild-type and endonuclease mutant L1 reporter constructs each mobilised very inefficiently in four cultured GBM cell lines.Here, using retrotransposon capture sequencing (RC-seq), we surveyed L1 mutations in 14 tumours classified as glioblastoma multiforme (GBM) or as a lower grade glioma. In four GBM tumours, we characterised one probable endonuclease-independent L1 insertion, two L1-associated rearrangements and one likely These experiments altogether highlight the consistent absence of canonical L1 retrotransposition in GBM tumours and cultured cell lines, as well as atypical L1-associated sequence rearrangements following DNA damage in vivo.The online version of this article (doi:10.1186/s13100-016-0076-6) contains supplementary material, which is available to authorized users. Ninety-ary GBM) . Primaryary GBM) . To dateary GBM) –6. Defecary GBM) , 8. DespAlu retrotransposons, in trans [Alu elements can also participate in DNA rearrangements driven by recombination [L1 retrotransposons are endogenous mutagens known to cause sporadic disease, including cancer . A full-in trans –27. L1 abination , 29. Altbination –38 and nbination –42, sevebination –32, 35.We hypothesised that L1-associated DNA rearrangements in GBM might occur via recombination or an atypical retrotransposition mechanism and therefore may lack the TPRT hallmarks required for L1 insertion recognition by previous genomic analyses. Alternatively, we considered that L1 insertions in GBM could be restricted to sub-clonal and highly heterogeneous events. We therefore applied deep retrotransposon capture sequencing (RC-seq) , 42 to 1Tissues were obtained from 14 patients undergoing surgical removal of a brain tumour at the Department of Clinical Neurosciences, Western General Hospital, Edinburgh, UK. All patients gave informed consent for tumour and peri-tumoural tissue removed in the normal course of surgery, and blood obtained intra-operatively, to be used for research. Ethical approval for the study was granted to P.M.B. by the East of Scotland Research Ethics Service (SR018). Tissue designation as ‘tumour’ or ‘adjacent brain’ was determined at the time of sampling based on pre-operative imaging, intra-operative image guidance and macroscopic inspection. Blood was sampled from 9 patients  tumour and adjacent brain from patients #1-#7, ii) tumour and adjacent brain from patients #8-#14 and iii) blood from patients #6-#12 and #14 were hybridised using a third generation (V3) RC-seq capture protocol involving only two optimised, custom locked nucleic acid (LNA) oligonucleotide probes  respectively targeting the 5′ and 3′ ends of L1.4  Fig. 1a1a. LNA phttps://github.com/adamewing/tebreak/tree/f7f01c1) with settings –mincluster 2, −minclip 30, −minq 1. Briefly, TEBreak aligned RC-seq reads against the human reference genome (hg19) using BWA-MEM [http://broadinstitute.github.io/picard/). Non-duplicate reads that aligned partially to the reference genome but had ≥30 nt soft-clipped from either end were retained; clipped ends were then aligned against L1.4 using the same BWA-MEM settings. Split-read mappings joining an L1 to the reference genome were then clustered and annotated for the presence of TSDs or deletions. Clusters with at least two reads supporting a consistent breakpoint were output in VCF format and further post-processed using the summary.py script included in the TEBreak distribution. This further filtered candidates by ensuring a consistent breakpoint between BWA-MEM and BLAT [RC-seq read data were analysed using TEBreak  following manufacturer’s instructions for total RNA isolation. Quantification was performed using NanoDrop 1000 . 2 μg total RNA was treated with DNase I  and used as template for cDNA synthesis with SuperScript III Reverse Transcriptase  following manufacturer’s instructions. 2 μg total RNA was processed as described with no reverse transcriptase added to the cDNA synthesis reaction for further use as negative control (RT-). cDNA from adjacent brain tissue was diluted to final concentrations of 1:5, 1:10, 1:20, 1:40 and 1:80 and used to generate a standard curve for each primer set. Samples were diluted to 1:20 final concentration for qRT-PCR. Real time PCRs were performed using SensiFast SYBR Lo-ROX kit  and run on a ViiA 7 Real-Time PCR System  with standard curve experiment analysis settings. Negative control qRT-PCRs were performed using water as template  and 2 μl of RT- reaction; no amplification was detected. MeCP2 isoforms (NM_004992) were assessed using 5′-GAGGCGAGGAGGAGAGAC and 5′-TGGTAGCTGGGATGTTAGGG as forward primers for isoforms 1 and 2, respectively, and 5′-GCAGAGTGGTGGGCTGAT as a common reverse primer to amplify 154 nt of isoform 1 and 161 nt of isoform 2. Additionally, 143 nt of the MeCP2 exon 4, present in both isoforms, were amplified using 5′-CAGAGGAGGCTCACTGGAGA as forward primer and 5′-GGCATGGAGGATGAAACAAT as reverse primer. EGFR (NM_005228) was amplified on the 5′UTR (amplicon of 156 nt) and the junction between exons 11 and 12 (176 nt) with the following primers:5′-ACAGCTTTGAAGAGAGCAGTGGTT, 5′-AGTCTGCCCGTTCTCAGATCTL1 ORF2: 5′-TGCGGAGAAATAGGAACACTTTT, 5′-TGAGGAATCGCCACACTGACTL1 (L1.4) expression was assessed by targeting 61 nt of the 5′UTR and 85 nt of the ORF2 with the primers: L1 5′UTR:TBP, 5′-GCAAGGGTTTCTGGTTTGCC, 5′-GGGTCAGTCCAGTGCCATAAACTB, 5′-AGAAAATCTGGCACCACACC, 5′-TAGCACAGCCTGGATAGCAA156nt of TATA-binding protein mRNA  and 173 nt of beta actin mRNA  were amplified with the following primers:2) are as follows: MeCP2 isoform 1: −3.312, 29.329, 0.95; MeCP2 isoform 2: −2.946, 29.578, 0.943; MeCP2 exon 4: −3.796, 27.991, 0.897; EGFR 5′UTR: −4.057, 30.028, 0.931; EGFR exon 11–12 junction: −4.660, 28.317, 0.985; TBP: −3.754, 27.208, 0.973; L1 5′UTR: −3.094, 21.209, 0.996; L1 ORF2: −3.11, 18.836, 0.9992; ACTB: −3.954, 22.606, 0.999.Standard curve parameters for qRT-PCR  or ACTB (for L1). Statistical analysis was performed with Prism5 (GraphPad Software), applying a cDNA from patient #2 tumour was used as template for RT-PCR to amplify possible L1-MeCP2 chimeric transcripts. The following chimeric variants were tested; Exon 1–L1, using a forward primer within exon 1 5′-GAGGCGAGGAGGAGAGAC and a reverse primer within the new L1, 5′-CACCAGCATGGCACATGTAT. Exon 2–L1, forward primer (5′-TGGTAGCTGGGATGTTAGGG) in exon 2 and reverse primer within L1 (5′-CACCAGCATGGCACATGTAT). L1-Exon 3, with the L1 primer as a reverse primer (5′-GCACATTGTGCAGGTTAGTTAC) and a forward primer within exon 3, (5′-GCAGAGTGGTGGGCTGAT).5 ng of genomic DNA from patient #2 adjacent brain and tumour were used as a template for a PCR to amplify the deleted region within MeCp2 using 5′-AAATTAGCCAGGCGTGGTG as forward primer within the deleted region and 5′-TCCTGTTTTGTCTTACGTCTTGA as reverse primer downstream of the deleted region. The PCR conditions were as follow; 1U MyTaq DNA polymerase , 1× MyTaq Reaction Buffer, 2 μM primers and 5 ng template DNA in a 25 μL reaction volume with the following cycling conditions: 3 min at 92 °C, then 35 cycles of 15 s at 92 °C, 15 s at 56 °C and 15 s at 72 °C, followed by a single extension step at 72 °C for 10 min. PCR amplicons were resolved in a 2 % agarose gel and scanned . Amplicons present on the scanned picture was quantified using Image Studio Lite version 4 software (LI-COR Biosciences). Values were corrected for background and normalised to adjacent brain value.200 ng of genomic DNA extracted from tumour and adjacent brain tissues was bisulfite converted using EZ DNA Methylation-Lightning Kit  following manufacturer’s instructions. After purification, 2 μL was used as template for a PCR reaction using L1-Bis-F and L1-Bis-R primers as described by Shukla et al. . PCR reaIllumina libraries (TruSeq Nano DNA sample preparation kit) were generated from patient #2 tumour and adjacent brain and, for patient #8, tumour and blood genomic DNA. Libraries had an insert size of ~300 nt and were sequenced on an Illumina HiSeq X Ten platform . Reads were aligned to hg19 using BWA-MEM (parameters –Y –M –R < read group name>) and sorted using SAMtools . PCR dupGBM cell lines were purchased from American Type Culture Collection  and grown in a humidified, 5 % CO2 incubator at 37 °C in the complete media as described by the provider. DBTRG-05MG  cells were grown in ATCC-formulated RMPI-1640 medium  supplemented with 10 % foetal bovine calf serum , non-essential amino acids  and 100 U/mL penicillin, 0.1 mg/mL streptomycin  to generate complete media. M059J  cells were grown in media containing a 1:1 mixture of Dulbecco’s Modified Eagle’s Medium and Ham’s F12 Medium  supplemented with 10 % foetal bovine calf serum, non-essential amino acids and 100 U/mL penicillin, 0.1 mg/mL streptomycin. LN18  and LN229  were grown in Dulbecco’s Modified Eagle’s Medium  supplemented with 5 % foetal bovine calf serum and 100 U/mL penicillin, 0.1 mg/mL streptomycin. HeLa cells were grown in a humidified, 5 % CO2 incubator at 37 °C in the complete media, Dulbecco’s Modified Eagle’s Medium  supplemented with 10 % foetal bovine calf serum and 100 U/mL penicillin, 0.1 mg/mL streptomycin.NotI-Hf and BstZ17I restriction enzymes  at 37 °C for 2 h to obtain L1.3-D205A and the JJ-NotI-Hf/BstZ17I backbone. This backbone was also treated with alkaline phosphatase  for 30 min at 37 °C to use for subsequent cloning. Purified L1.3-D205A was ligated to the JJ-NotI-Hf/BstZ17I backbone using T4 ligase  for 2 h at room temperature generating JJ-L1.3-D205A. To obtain wild-type L1.3 (L1.3 WT) [EcoRI , NotI and BstZ17I simultaneously (2 h at 37 °C). Fragments corresponding to L1.3-NotI/EcoRI, L1.3-D205A-NotI/EcoRI, L1.3-EcoRI/BstZ17I and L1.3-D702A-EcoRI/BstZ17I were gel purified. L1.3-NotI/EcoRI and L1.3-EcoRI/BstZ17I were ligated to JJ-NotI-Hf/BstZ17I backbone to obtain JJ-L1.3 plasmid. JJ-L1.3-D205A-D702A was generated by ligation of L1.3-D205A-NotI/EcoRI and L1.3-D702A-EcoRI/BstZ17I to the JJ-NotI-Hf/BstZ17I backbone. Ligations were transformed in One Shot TOP10 Chemically Competent E. coli bacteria , and plated in LB-agar  and 100 μg/ml ampicillin . All plasmid sequences were confirmed by capillary sequencing.Plasmids carrying i) an L1 EN mutant (pCEP4-L1.3D205A)  and ii) L1.3 WT)  and L1.32O to 0.5 μg/μL. GBM cells were seeded in 6-well dishes in their respective complete media to ~25 % confluence. Cells were transfected at the time of seeding with FuGENE HD transfection reagent  following the manufacturer’s protocol using 1:4 DNA: FuGENE ratio. Each transfection well received 1 μg plasmid DNA, 4 μL FuGENE reagent and 2 mL of complete media. Media was changed 24 h post transfection and selection with blasticidin S HCl  began 4 days post transfection. Cells were selected for antibiotic resistance for 10 days using a final concentration of 5 μg/ml for HeLa, DBTRG-05MG and LN229 and 2 μg/mL for LN18 and M059J.13-14 days post transfection, cells were washed twice with 1xPBS and fixed and stained as described by Moran et al. [Plasmid DNA was purified on maxiprep columns  and diluted in sterile dHTransfection efficiency for each plasmid was calculated by flow cytometry . GBM cell lines were co-transfected as described above with 0.5 μg of each JJ construct and 0.5 μg of pCAG-GFP (plasmid that constitutively expresses GFP). 72 h post-transfection, cells were harvested and re-suspended in 1× PBS. Propidium iodide  was added to the samples for identification of dead cells. GFP positive cells were gated based on fluorescence of untransfected cells and transfection efficiency calculated as the percentage of GFP positive cells using FlowJo 10.0.8 software   gene by RC-seq that was confirmed by PCR  reductions in tumoural expression of both of the two main MeCP2 transcript isoforms  L1 mRNA abundance in patient #2 tumour versus adjacent brain   Fig. . Replicap < 0.0001, t-test)  Fig. , a majorst) Fig. . As EGFRst) Fig. . Given tTo place the PCR validated L1-associated mutations in MeCP2 and EGFR into a broader context of genomic abnormality, we analysed WGS data from patients #2 and #8 (Additional file As noted above, the tumour sample from patient #8 showed a remarkable amplification of EGFR (>50 fold, Fig. To assess whether GBM cell lines support canonical or ENi L1 retrotransposition in vitro we performed an established cultured cell retrotransposition assay  on HeLa These experiments reveal rare L1-associated mutations caused by recombination or L1 ENi retrotransposition in GBM tumours, accompanied by an absence of TPRT-driven L1 insertions. Endonuclease-independent L1 insertions have been reported by several prior studies employing engineered L1 reporter constructs in cultured cancer cells or cells otherwise deficient for DNA damage repair factors , 72, 73,Alu:Alu recombination. Notably, the Alu-flanked L1PA2 donor sequence, located on chromosome 9, was 5′ truncated and did not encode a viable L1 ORF2p. The involvement of an older L1 family in a tumour-specific DNA rearrangement event is reminiscent of one of the earliest L1 mutations detected in cancer, in that case affecting the myc locus in breast tumour [Alu during ENi L1 mobilisation in trans, in agreement with a previous observation where 71 % of detected ENi L1 mobilisation events involved non-L1 DNA [Although identified at first by RC-seq as a potential L1 mobilisation event, further characterisation of the MeCP2 L1 mutation indicated a probable DNA rearrangement event mediated by t tumour . A lessen-L1 DNA , and invn-L1 DNA , 73, 75.Notably, the EGFR L1 mutation provides an interesting example because, despite not having a direct effect on gene activity, it is one of very few tumour-specific L1 mutations noted in a major oncogene or tumour suppressor since the first such example was discovered more than 20 years ago by Miki et al. .de novo L1 insertions in this context is intriguing, given frequent somatic L1 mobilisation via TPRT in neural cells [Previous studies employing high-throughput sequencing reported no tumour-specific L1 insertions in brain tumours –32, 35. al cells –42, inclal cells . This coAlthough we conclusively find that L1 mobilisation is a rare event in GBM, our discovery of atypical L1-associated mutations in MeCP2 and EGFR demonstrates that L1 can otherwise contribute to GBM genome abnormality in key loci regulating neural cell differentiation and proliferation. Future experiments are required to ascertain whether this phenomena correlates with patient prognosis and whether potential DNA damage caused by chemotherapy or radiotherapy  activate


Programmed death ligand (PD-L1) expression was associated with tumor immune escape and subsequent poor prognosis in non-small cell lung cancer (NSCLC). This expression was higher in patients with EGFR-mutated NSCLC tumors than in those with EGFR-wild-type (WT) NSCLC tumors. We therefore hypothesized that poor prognosis mediated by higher PD-L1 may be partially through conferring resistance to tyrosine kinase inhibitor (TKI) in NSCLC regardless of EGFR mutation. The change in PD-L1 expression following gene manipulation corresponded with changes in expression of HIF-1α and YAP1. The expression of HIF-1α and YAP1 was concomitantly decreased by PD-L1 silencing or by ROS scavenger treatment ; however, a ROS inducer treatment (pyocyanin) completely reversed the decreased expression of both genes in EGFR-mutated and -wild-type (WT) NSCLC cells. The MTT assay indicated that the inhibitory concentration of gefitinib yielding 50% cell viability (IC50) depended on PD-L1-mediated YAP1 expression. Mechanistic studies indicated that upregulation of YAP1 by PD-L1 might be responsible for EGFR mutation-independent TKI resistance via the ROS/HIF-1α axis. An unfavorable TKI response was more common in patient tumors with high PD-L1 or YAP1 mRNA expression than in patient tumors with low mRNA expression of these genes. In conclusion, PD-L1 might confer EGFR mutation-independent TKI resistance in NSCLC cells via upregulation of YAP1 expression. The binding of programmed death ligand 1 (PD-L1) to its receptor PD-1 leads to T cell apoptosis, which, in turn, triggers tumor immune surveillance . PD-L1 oTumor metastasis and drug resistance represent a crucial limitation in human cancer therapy. EGFR-tyrosine kinase inhibitors (EGFR-TKIs) have known clinical benefits in the treatment of EGFR-mutated NSCLC when compared to EGFR-WT NSCLC , 7. UnfoEGFR-TKIs down-regulate PD-L1 expression in EGFR-mutated NSCLC through inhibition of the NF-κB signaling pathway . PD-L1 eThe development of the hypoxic tumor microenvironment, which is associated with tumor progression and metastasis, depends on the presence of myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs) , 24, andOur preliminary data showed that PD-L1, HIF-1α, and YAP1 expressed concomitantly in EGFR-mutated and EGFR-WT NSCLC cells. YAP1 confers chemoresistance in NSCLC cells , and YAPFour EGFR-mutated and EGFR-WT cell types were collected to verify whether PD-L1 expression could be associated with HIF-1α and YAP1 expression. Western blotting indicated a higher PD-L1 expression in EGFR-mutated cells than in EGFR-WT cells Figure . This exWe next examined the possibility that PD-L1 could be responsible for TKI resistance in NSCLC cells. Two EGFR-mutated and EGFR-WT cell types collected for manipulation of PD-L1 expression to test this possibility. The MTT assay indicated that the inhibitory concentration of gefitinib yielding 50% cell viability (IC50) decreased markedly by PD-L1 silencing in H1650, H1975, and CL1-5 cells. The IC50 values were gradually increased by ectopic PD-L1 expression in CL1-0 cells Figure . These rWe next examined whether PD-L1 may induce HIF-1α expression through ROS generation, thereby upregulating YAP1 expression at the transcription level. H1975 and CL1-0 cells were collected to manipulate PD-L1 expression and/or treatment with a ROS inducer (pyocyanin) or a ROS scavenger . Western blotting showed a concomitant decrease in expression of HIF-1α and YAP1 following shPD-L1 transfection or treatment with NAC. However, the decrease in HIF-1α and YAP1 expression by PD-L1 silencing reversed by pyocyanin treatment in both cell types , YAP1 (verteporfin), and ERK (U0126), when compared to both cell types without these inhibitor treatments  between 1998 and 2004. Patients were asked to submit written informed consent; the study was approved by the Institutional Review Board (TMUH No. 201301051). The tumor type and stage of each collected specimen were histologically determined in accordance with the World Health Organization classification system. Among these patients, 46 patients were received TKI therapy when these patients were occurred tumor relapse. The tumor response to TKI therapy collected from chart review.TL-1 cells was kindly provided by Dr. Y.-W. Cheng  . H1650, The expression of PD-L1 mRNA and YAP1 mRNA levels in patients’ tumors was determined by real-time PCR analysis as described previously .shRNA was purchased from the National RNAi Core Facility, Academia Sinica, Taiwan. Plasmids containing the PD-L1 expression construct were constructed by cloning the full-length human PD-L1 cDNA (GenBank accession number NM_014143) into the pcDNA3.1 eukaryotic expression vector, which also expresses a neomycin (Neo) resistance gene. YAP1 and HIF1 overexpression plasmids were provided from Origene . The transfection and stable clone selection procedures have been described previously . All datin vitro cytotoxic effects of these treatments were determined by MTT assay (at 570 nm). The data were obtained from three independent experiments.The cell lines were cultured in 96-well flat-bottomed microtiter plates supplemented with RPMI 1640 and DMEM containing 10% heat-inactivated fetal bovine serum, 100 units/mL penicillin, and 100 units/mL streptomycin in a humidified atmosphere containing 95% air and 5% CO2 at 37°C in a humidified incubator. Before TKI treatment , the cells cultured in the exponential growth phase were pretreated with shRNAs, PD-L1 and BAG-1 overexpression plasmid for 24 h. After 48 h incubation, the 2) at a final cell density of 1 to 2*106 cells/mL, 100 μL of a single-cell suspension (1-2*105 cells) was incubated with 5 μL Annexin-V–FITC and 5 μL propidium iodide (PI) for 15 minutes at room temperature in the dark. After addition of 400 Ml of binding buffer, the samples were analyzed with a BD FACS Calibur flow cytometer (BD Biosciences) within 1 hour. For each sample, 10,000 events were counted.The cells were collected by trypsinization and centrifugation at 1,000 g for 5 minutes. Following resuspension in binding buffer .


Unfortunately, after publication of this article , it was The original article has also been updated.Figure 1 Binding and blocking properties of LY3300054. Panels a–c: 96-well plates were coated with recombinant human (a), cynomolgus (b), or murine (c) PD-L1-Fc fusion protein (100 ng/well each). Bound LY3300054 was detected using HRP-conjugated anti-human Fab antibody and addition of chromogenic substrate (OD at 450 nm). 96-well plates were coated with 100 ng/well of recombinant PD-1 (d) or B7-1 protein (e), then incubated with a mixture of biotin-conjugated PD-L1 and either LY3300054 or human IgG1 antibodies. Plate bound PD-L1 was detected using HRP-conjugated streptavidin and addition of chromogenic substrate (OD at 450 nm). In all experiments, each data point is the average of two replicates. Data (a-e) are representative of multiple independent experimentsFigure 2 Identification of LY3300054 epitope residues in human PD-L1. Panel a: CLUSTALW multiple sequence alignment of domain 1 of human (hu), canine (ca), and murine (mu) PD-L1 and hu-PD-L2 to identify the LY3300054 species specificity anchors on hu-PD-L1. Underlined is the human PD-1 6 Å binding site on hu-PD-L1 (according to PDB: 4ZQK (26602187)). An alignment position is marked with (*) if both mu-PD-L1 and ca-PD-L1 substitutions differ from the hu-PD-L1 sequence. An alignment position is marked with (:) if either the mu-PD-L1 or ca-PD-L1 substitution differs from the hu-PD-L1 sequence. Panel b: Position N63 on human PD-L1 is a specificity anchor for LY3300054. Canine-to-human mutation K63 N (▲) rescues the ELISA binding of LY3300054 to canine PD-L1. Like wild type ca-PD-L1-Fc (●), canine-to-human mutant N69H (△) does not bind LY3300054


We synthesized Ag nanoparticle-decorated multilayered graphene nanosheets (Ag-graphene) from graphite nanoplatelets and silver nitrate through 90–100 s of microwave exposure, without the use of any mineral acids or harsh reducing agents. Fe nanoparticle-decorated carbon nanotubes (Fe-CNTs) were grown on polypyrrole (PPy) deposited on woven Kevlar fibre (WKF), using ferrocene as a catalyst, under microwave irradiation. Fe-CNTs grown on WKF and Ag-graphene dispersed in polyester resin (PES) were combined to fabricate Ag-graphene/Fe-CNT/PPy-coated WKF/PES composites by vacuum-assisted resin transfer moulding. The combined effect of Fe-CNTs and Ag-graphene in the resulting composites resulted in a remarkable enhancement of tensile properties (a 192.56% increase in strength and 100.64% increase in modulus) as well as impact resistance (a 116.33% increase). The electrical conductivity significantly increased for Ag-graphene/Fe-CNT/PPy-coated WKF/PES composites. The effectiveness of electromagnetic interference shielding, which relies strongly on the Ag-graphene content in the composites, was 25 times higher in Ag-graphene/Fe-CNT/PPy-coated WKF/PES than in neat WKF/PES composites. The current work offers a novel route for fabricating highly promising, cost effective WKF/PES composites through microwave-assisted synthesis of Fe-CNTs and Ag-graphene. As most electronics now-a-days are wireless, electromagnetic interference (EMI) has been receiving increasing attention due to its impact on EMI-sensitive electronic devices and telecommunication. Shielding of these unnecessary electromagnetic (EM) waves has become a big concern for many applications, including those in military and commercial sectors157810132SO4, KMnO4Graphene, a unique and very promising material, is a one-atom thick 2-D aromatic sheet consisting of sp15Kevlar is notable for its high strength, tenacity, and impact resistance, and has major applications as a body and vehicle armour material, as well as a structural composite material in automobiles and aircraftsin-situ polymerization of pyrrole. Fe nanoparticle-decorated CNTs (Fe-CNTs) were grown on the surface of the PPy-deposited WKF within a very short period of time (15–30 s) via a unique microwave-assisted method. The conducting layer of PPy on the surface of insulating WKF helps with the rapid increase of the temperature above 1100 °C to create arcs and sparks when exposed to microwave radiation. Ag nanoparticle-decorated multilayered graphene nanosheets (Ag-graphene) were also synthesized separately from graphite nanoplatelets (XGnP) by a similar eco-friendly microwave technique. Composites of polyester resin (PES) with Fe-CNTs grown WKF were prepared through a vacuum-assisted resin transfer moulding (VARTM) technique. Prior to the VARTM process, Ag nanoparticle-decorated graphene layers were dispersed in the resin by sonication. The schematic diagram for the preparation of the composites is shown in Hereby, the present study focuses on the combined effect of Fe-CNT and Ag-graphene on the tensile properties, impact response, electrical conductivity and EMI shielding effectiveness of the composites. Polypyrrole (PPy) was initially, deposited on the surface of a woven Kevlar fibre (WKF) sheet through The scanning electron microscopy (SEM) micrographs of stacked layers of XGnP, the precursor for the formation of graphene nanosheets, are shown in in-situ PPy-deposited WKF had a rough and uneven surface (5H5) were formed because of the rapid rise in temperature. The formed Fe atoms might be in a molten state that may later pave the way for the formation of Fe nanoparticles through condensation of the molten Fe atoms. The sources of C required for the CNT growth were cyclopentadienyl, toluene and hexane. The C atoms remained dispersed across the Fe nanoparticles above the substrate. Graphitic walls, which later commenced the CNT growth, formed through arrangement of C atoms that precipitated from the Fe nanoparticlesUncoated WKF had a uniform, smooth surface , whereas surface , with PP surface . The PPyThe transmission electron micrographs (TEM) of Fe-CNT and Ag-graphene are shown in −1. This D band, a result of the one phonon lattice vibration process, is generally very weak in the spectra of both graphite and graphene. If it is intense, it indicates that the synthesized material contains many defects. Thus, the amount of defects present in graphene is directly proportional to the intensity of the D bandG/I2D) differentiates between less exfoliated graphite and few-layered graphene layersG/I2D decreased for multilayered graphene indicating good exfoliation of graphite layers. The peak shift in graphene was a result of interactions between the stacked graphene layers, which tends to shift bands to lower wave numbers. The intensity of the 2D band was less pronounced compared to single layer graphene nanosheets but higher than that of XGnP. The 2D band was composed of two peaks in the XGnP spectra of XGnP (2D-1 and 2D-2), as shown in 3031293031−1 due to quinoniod bipolaron and polaron structures of PPy, and peaks at 1061 and 1111 cm−1 associated with –CH stretching−1 in the spectrum of neat WKF increased considerably in intensity after growth of Fe-CNTs on the surface of WKF due to the D and G bands of CNT, while a new peak appeared at 2665 cm−1 corresponding to the G′ band.et al. also synthesized Ag nanoparticles decorated graphene and reported a similar finding in the XRD analysis of the spectra3C), 44.83° ((110) plane of Fe), 55.81° ((004) plane of CNT), and 76.61°((110) plane of CNT) in the spectrum of Fe-CNT grown PPy-coated WKF, in combination with the increased intensity of the peaks at 2θ = 20.94°and 23.98° confirmed the growth of Fe-CNTs on the surface of PPy-coated WKFThe X-ray diffraction spectra of XGnP and Ag nanoparticle-decorated graphene are shown in The stress-strain curves, tensile strengths and moduli of the WKF composites, to evaluate the degree of reinforcement provided by Fe-CNTs and Ag-graphene, are shown in et al. previously reported an improved impact resistance after the incorporation of MWCNTs into woven Kevlar/epoxy compositeset al. reported improvement in mechanical properties after incorporation of Ni nanoparticles decorated graphene nanoplatelets in Cu matrix compositeThe impact resistance of the WKF composites was determined from the plot of absorbed energy vs. time as depicted in The velocity-time response curves of the composites are presented in et al. prepared PPy-coated fly ash, exposed it to microwave radiation to synthesize CNT, and reported that CNT improved the electrical conductivityThe electrical conductivity values of the WKF composites are shown in R), absorption (SEA) and multiple-reflection (SEM) SEs of EM radiation incident on the surface of a material. The impedance between the shielding material and air is related to the reflection, the energy dissipation of absorbed incident EM waves resulted in absorption, and the non-uniformity of the interior of the shielding composite panel resulted in multiple reflection. The multiple reflection is taken onto account only in case of porous or foam material where there is large surface areas47The total EMI shielding effectiveness (SE) of each WKF composite is revealed in μ, σ and ε denote the magnetic permeability, electrical conductivity and permittivity of the medium, respectively, and ω and f represent the angular frequency of the radiation and frequency of the electromagnetic wave, respectively. From the above equations, it is clear that electrical conductivity is one of the main parameters for reflection suppression and a major component in shielding by absorptionwhere Here, the thickness of all the samples was 0.35 mm. Since the fabricated composites were very thin and prepared with only a single layer of plain weave WKF, low SE values were obtained for all of them. The SE value at 0.20 dB was the lowest for neat WKF/PES composites. Deposition of conducting PPy polymer on WKF resulted in a higher total EMI SE for PPy-coated WKF/PES composites than for neat WKF/PES composites. The Fe-CNTs synthesized on the surface of PPy-coated WKF had a significant effect, enhancing the EMI SE of Fe-CNT/PPy-coated WKF/PES composites. This was mainly ascribed to the arrangement of the conducting interconnected networks by Fe-CNTs in the PPy-coated WKF/PES composites. The interrelated networks of CNTs and ferromagnetic Fe nanoparticles in the composite interacted with the incident EM radiation, thereby acting as a shield and leading to the enhanced SEin-situ polymerized on the surface of WKF. The direct synthesis of Fe-CNTs on the fibres prevented the problem of agglomeration. Scanning electron microscopy showed the growth of CNTs with Fe nanoparticles at their tips on the surface of WKF, as well as the decoration of multilayered graphene nanosheets with Ag nanoparticles. The Fe-CNTs and Ag-graphene were further characterized by Raman spectroscopy and X-ray diffraction analysis. The tensile strength and modulus of PPy-coated WKF/PES composites was enhanced by the growth of Fe-CNTs on their surface, while an increase in the amount of Ag-graphene further enhanced the mechanical properties of the composites. The highest increase in tensile strength (192.56%) and modulus (100.64%) was achieved with 1% Ag-graphene in the composites. A significant improvement in absorbed impact energy and penetration limit of the Ag-graphene/Fe-CNT/PPy-coated WKF/PES compared to neat WKF/PES was observed due to the synergistic reinforcing effects of Fe-CNTs and Ag-graphene. Fe-CNT/PPy-coated WKF/PES composites with 1% Ag-graphene displayed the highest impact resistance and an absorbed energy increase of 116.33%. The in-situ polymerization of pyrrole induced some conductivity in WKF/PES, but the combined effect of Fe-CNTs and Ag-graphene increased it to a remarkable extent. Finally, the total electromagnetic interference (EMI) shielding effectiveness (SE) value of a single sheet of neat WKF/PES was 0.20 dB, butwas25-fold higher for Fe-CNT/PPy-coated WKF/PES composites containing 1% Ag-graphene. The higher the Ag-graphene content, the higher the total EMI SE of the composites. Although an increase in EMI shielding properties was observed, more studies are required to investigate the nanofiller effects for thick composites prepared with multilayered sheets of WKF. Nonetheless, the improved interaction and formation of a conductive network between the WKF, PPy, Fe-CNT, Ag-graphene and PES in the composites contributed to their enhanced mechanical, electrical and EMI shielding properties in this study.Fe nanoparticle-decorated carbon nanotubes (Fe-CNT) were grown on polypyrrole (PPy) deposited on woven Kevlar fibre (WKF), and composites of the Fe-CNT grown WKF and Ag-graphene dispersed in polyester resin (PES) were then fabricated by a vacuum-assisted resin transfer moulding technique. A microwave-induced technique was utilized for the synthesis of both Fe-CNTs and Ag nanoparticle-decorated multilayered graphene nanosheets, which is a very economical and time saving approach. WKF was surface functionalized and PPy was 3.6H2O), ferrocene, hexane and toluene were purchased from Sigma-Aldrich . Duksan pure chemicals, Korea, supplied reagent grade hydrogen peroxide. PES (LSP-8020B) and methyl ethyl ketone peroxide were obtained from CCP Composites and ARKEMA, Korea. Acetone and ethanol was supplied by J.T. Baker  and used as received.Woven Kevlar fibres (WKF) (Kevlar 49) were obtained from JMC Co., Ltd. . Microwave Research & Applications Inc. USA (BP110 laboratory grade microwave oven) supplied a laboratory scale microwave oven. Silver nitrate was obtained from Daejung Chemicals and Metals Co. Ltd. Korea and graphite nanoplatelets from Hanwha Nanotech Corp, Korea. Pyrrole, ferric chloride hexahydrate (FeCl3.6H2O (12% w/v), which was used as the oxidizing agent. WKF samples were taken out of the FeCl3.6H2O solution after 15 min for complete polymerization of pyrrole. Finally, PPy-coated WKF samples were washed several times with deionized water and dried overnight in an oven at 60 °C.Methanol and water were mixed in a 50:50 ratio and 12% v/v pyrrole was added. WKF samples were soaked in the pyrrole solution for 60 min at room temperature and then placed in a sonicator bath. The samples were taken out after 60 min and put in an aqueous solution of FeClA ferrocene solution was prepared by dissolving ferrocene in toluene, and the PPy-coated WKF samples were placed in this solution at a 1:1 ratio of sample to ferrocene. The samples were then placed in a sonicator bath for 15 min for good distribution of ferrocene throughout the samples. The growth of Fe-CNTs on the surface of the fibres was enhanced by the addition of 0.25 mL of hexane. After leaving the samples in air for 1 min to let the solvents evaporate, they were exposed to microwave irradiation.Natural graphite, hydrogen peroxide and ammonium persulfate were mixed in a glass bottle in a 2:1:0.1 weight ratio, and exposed to microwave irradiation for 30 s. Then, urea and ammonia were added as reducing agents, and 20 mL of 1 N silver nitrate was added to the mixture as the metal precursor. The mixture was then sonicated for 1 h and subjected to microwave irradiation for 90–100 s. The precursors exfoliated rapidly, followed by some arc and lightning on exposure to microwave irradiation. The resulting Ag-graphene was dispersed in unsaturated PES by sonication before the preparation of the composites through VARTMA Nova Nano SEM 230  was operated at 15 kV to investigate the growth of Fe-CNTs on the surface of WKF and Ag-graphene. Normal TEM  was used to characterize the growth of Fe-CNT and Ag-graphene. A micro-Raman Microscope (WITec) was used using 532 nm excitation via the back scattering geometry with the polarizer and analyzer parallel to one another to obtain the Raman spectra. The X-ray diffraction patterns of the samples were studied using a wide-angle X-ray diffractometer  at a 40 kV operating voltage with a 20 mA current using crystal-monochromated Cu–Kα radiation in the range of 5°–80° (2θ). Tensile tests for the samples were performed with an Instron 5982 universal testing machine at a displacement rate of 2 mm/min with a maximum load of 100 kN following ASTM D3039 standard. The results for each composite were an average of three specimens. The impact test was carried out using a drop-weight impact tester  in accordance with the ASTM D5628-10 standard. The circular clamp was loaded with a 5-kg impactor with a 40 mm diameter, and data were collected between the initial impact contact and penetration points. 200 J of energy was applied to all the samples to fully perforate them. Electrical conductivity tests were performed with a 6517 multimeter . The EMI SEs of the composites were measured using an EMI SE test system . The tests were carried out in accordance with ASTMD4935, utilizing a coaxial cable with type-N connectors. The frequency was measured up to 1 GHz, and the attenuation of EM radiation was obtained as a function of frequency.How to cite this article: Hazarika, A. et al. Microwave-synthesized freestanding iron-carbon nanotubes on polyester composites of woven Kevlar fibre and silver nanoparticle-decorated graphene. Sci. Rep.7, 40386; doi: 10.1038/srep40386 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


In this paper, we investigated the effect of humidity on paper substrates and propose a simple and low-cost method for their passivation using ZnO nanoparticles. To this end, we built paper-based microdevices based on an interdigitated electrode (IDE) configuration by means of a mask-less laser patterning method on simple commercial printing papers. Initial resistive measurements indicate that a paper substrate with a porous surface can be used as a cost-effective, sensitive and disposable humidity sensor in the 20% to 70% relative humidity (RH) range. Successive spin-coated layers of ZnO nanoparticles then, control the effect of humidity. Using this approach, the sensors become passive to relative humidity changes, paving the way to the development of ZnO-based gas sensors on paper substrates insensitive to humidity. Cellulose fiber-based paper has been used for long time in missives, flyers, books or newspapers, because it is both chemically and mechanically stable under atmospheric conditions and absorbs ink readily. In the present-day, paper can be also used as substrate for the fabrication of electronic devices such as antennas, sensors, photovoltaics or energy storage devices ,2,3,4. CPaper is a good candidate for gas-sensing applications thanks to its inherent porous structure and rough surface, which provides a large surface area compared to flat glass or a nonporous plastic substrate . SeveralPaper is also the perfect candidate for low-cost and disposable humidity sensors due to its capacity to absorb water vapor. However, only a few concrete examples of humidity sensors using paper substrate as the sensing element can be found in the literature. For instance, a fully printed chip-less radio frequency identification sensor for humidity monitoring applications was reported in . AnotherAlthough paper can be a promising candidate for humidity sensing, its use for other gas sensors can be restricted by the same effect, affecting the gas sensor’s selectivity and influencing their performances. Many studies have focused on devising methods for improving the surface hydrophobicity of paper, such as laser ablation to modify the surface morphology and/or change the surface energy  and plasIn this context, we propose a simple, effective and low-cost approach to study and reduce the effect of humidity on the electrical properties of simple printing papers with different morphology. As a first step, we fabricated paper-based microdevices by laser ablating metal layers deposited on the surface of the paper substrates and characterized them in detail under known relative humidity levels at room temperature. This enabled us to explore the role of surface morphology in the ability of a particular paper to act as a humidity sensor.As a second step, we examined a time- and cost-efficient methodology—inspired by Nature’s approach to exploit nanoparticle formations for functionality enhancement —as a pot−2 basis weight and a glossy, photographic quality paper of a 200 gm−2 basis weight. Interdigitated electrodes (IDEs) were directly patterned on them using an inexpensive and fast fabrication process, which does not require the high-cost semiconductor manufacturing equipment normally used for silicon microfabrication. The process sequence is schematically illustrated in Even though the most popular paper used in sensor technology is the Whatman no.1 chromatography paper , this work focuses on two other paper substrates: a plain printing paper of a 80 gm2 rectangular pieces 2.2H2O, Merck Millipore, Darmstadt, Germany) into analytical grade ethanol  at a concentration of 40 mM [In our work, sol-gel solutions were prepared by dissolving zinc acetate dihydrate  their size distribution shown in In order to examine whether the successive coating with the ZnO sol-gel results in a uniform film, a patterned silicon wafer with IDE electrodes was deposited with ZnO nanoparticles under the same conditions . The SEM image of the resulting film b shows tA custom-designed experimental setup was developed in order to perform the measurements in a controlled environment. The setup consists of a sealed Teflon chamber, inside which the sample is carefully placed, along with the sensing tip of a portable hydrometer, in order to measure the changes in the RH in real-time. Mass-flow controllers and flow meters from Brooks Instruments  were used to determine the actual flow rates and concentration of gases. Nitrogen was used as a carrier gas for the water vapors. The gas was driven inside a sealed glass bottle containing water (bubbler), which caused the formation of bubbles and the resulting vapors were then guided to the Teflon chamber where the device was located. Responses to relative humidity changes were recorded by measuring the drop in resistance across the IDEs, using an amperometer  driven by a custom-designed Labview-based interface. All measurements were performed in room temperature (T = 25 °C).The effect of humidity on paper largely depends on its surface morphology and composition, which in turn depend on the manufacturing process. A structural characterization of both types of paper is thus required in order to understand paper behavior under controlled humidity levels.3) introduced into the pulp during paper manufacturing in order to fill the pores of the cellulose matrix and improve paper quality, especially light scattering, ink absorbance and surface smoothness. Similar results were also obtained for the glossy paper. However, in this case, in addition to calcium and oxygen, silicon, as well as aluminum peaks were identified.Energy-dispersive X-ray spectroscopy (EDX) was used in order to analyze the composition of the various papers. Although the exact manufacturing conditions of the paper substrates used is not known, the presence of Si and Al is not unusual since it can be attributed to the specifics of the manufacturing processes for improving paper quality and characteristics. Dry-strength additives, or dry-strengthening agents, are chemicals that improve paper strength under normal conditions. These improve the paper’s compression strength, bursting strength, tensile breaking strength, and delamination resistance. Typical chemicals used include cationic starch and polyacrylamide (PAM) derivatives. These substances work by binding fibers, often with the aid of aluminum ions in paper sheet. In addition, Si is used during paper fabrication, a fact that could explain the EDX signal. SEM characterization of both paper substrates with patterned IDE was performed in order to determine the differences in morphology, as shown in −2 plain paper indicating a rough and porous surface morphology , as a function of humidity level for 80 and 200 gm−2 paper-based devices; the plain paper exhibits a higher response in the 20% to 50% RH rage than the glossy one.+ and OH−. Thus, the current can flow due to ionic conduction decreasing resistance along the IDE electrodes and resulting to a high sensing response for both papers. The dependence of electrical resistance on water diffusion through the porous structure of paper is also indicated by the difference in the response and recovery times obtained for the two paper substrates.Indeed, a porous and rough paper surface is more prone to water molecule absorption than a smooth one, since its surface area is larger. In fact, at moderate RH (40%), diffusion of water vapor further inside the volume of paper occurs through the network of air spaces among the cellulose fibers. As water progresses through the stack, an amount is sorbed onto the fibers’ surface, which results in water to lose its original concentration. The cellulose fibers absorb and desorb water into the capillary air space next to them, with little to no transmission of water molecules between them, therefore any further movement of wateroccurs almost entirely through air spaces. This absorption causes the paper resistance, as measured by the IDE electrodes, to decrease. At higher percentages of RH, water dissociation occurs on the moist cellulose fiber, creating HGenerally, a rough and porous surface is more capable of water absorption than a smooth one, which results to a faster response time. The recovery time follows the same trend as the response time with a faster dehumidification for the plain paper up to 60% RH as shown in The humidity sensing performance of both papers in terms of response/recovery time and sensitivity are summarized in The effect of humidity on paper can diminish its potential for use in electronic devices, therefore, this issue needs to be addressed. In order to eliminate the humidity effect, we propose an original solution consisting in the deposition of ZnO nanoparticles. Layers of ZnO nanoparticles were deposited using a standard and low-cost spin-coating method to allow the creation of a uniform layer on the entire surface of the paper.For both kinds of paper, resistance decreases as the number of successive spin coated layers increases. For the plain paper, a 30% reduction in resistance, from 1.96 MΩ to 1.55 MΩ, was observed after 10 deposited layers from a 40 mM sol-gel and the glossy paper exhibits a similar behavior, except for the first deposition. Upon deposition of a single ZnO layer, resistance increased compared to the uncoated paper. Subsequent layer deposition resulted in ever lower resistance, as can be seen in o/R), measured between the IDE electrodes, of the 80 gm−2 and 200 gm−2 paper-based devices as a function of the RH, after each spin-coating of ZnO nanoparticles. We observe that the device’s response to humidity decreases with the number of coatings. After five deposited layers, the effect of humidity on the paper is considerably decreased. These results reveal that the ZnO layer, at least under the experimental conditions used in this work, acts as a passivation layer to humidity prohibiting water molecules from interacting with the cellulose fibers. All measurements were conducted in fixed time intervals (30 min) for each %RH, where the device’s passivation remain unchanged for the duration of each measurement.−2 paper and results in the complete passivation of paper after 10 deposited layers. This can be attributed to the different morphology and fabrication process of the two different papers, as it can be seen from SEM images  like ethanol.