This nanoflower like architecture was decorated on 3D-graphene on Graphite electrode to design the solid-state-supercapacitor prototype device of dimensions of 23.6 × 22.4 × 0.6 mm3 having considerable high Csp of 58.0F/g and energy density of 24.59 Wh/Kg, and power density of 8.8 W/Kg. Four fabricated supercapacitors were connected in series for real state practical demonstration using the light emitting diode that remains enlightened for 40 s by charging it only for 25 s. This study demonstrates the 3D-graphene/MoS2 nanohybrid has a quite high overall potential window nearly about 2.7 V (−1.5 to +1.2 V) in KOH-PVA medium which can be used for the development of solid-state supercapacitors thereby completely eliminating the need for any expensive ionic liquid mediums thus building an exciting potential for high-performance energy storage/transfer devices.Portable and matured energy storage devices are in high demand for future flexible electronics. Flowery shaped MoS Due to the increase in demand for energy, the non-renewable sources like fossil fuels are continuously decreasing and will automatically come to an end in the near future. To meet this demand, alternate renewable energy sources has been projected like solar, wind and hydropower. However, a suitable storage device like Li-ion batteries and Na-air batteries and so on are currently lagging behind with a storage capacity of only 1% of the total renewable energy with longer charging time. Presently, batteries are used in wide variety not only for energy storage but also for the energy supply devices2. The use of a battery in large scale sophisticated electronic devices is limited due to its heavy weight, portability, flexibility and other issues4. Currently, researchers are looking for alternative energy storage devices that effectively store and provide energy as per requirement. Supercapacitors are portable, flexible5 and transparent unlike conventional batteries that draw the attention of the researchers to extract its interesting properties. Along with these physical advantages, supercapacitor has high charge storage capacity, high power density, low energy density, longlife time and short charging time as compared to traditional batteries8. A notable improvement in performance has been achieved through recent advances in understanding charge storage mechanisms and the development of advanced nanostructured materials in supercapacitor devices9. Supercapacitors are classified mainly in two categories such as, pseudocapacitor and electrical double layer capacitance (EDLC)8. In case of pseudo capacitor, the electrical energy is stored via faradaic reaction in electrolyte and electro-active species on the electrode surface. There are several reports in the literature based on pseudocapacitors made of various nanomaterials as well as their composite, such as SnO2-V2O5, Cobalt pyrophosphate, Metal oxide-CNT, RGO-metal sulfide, MnO2-veritcally aligned graphene, etc. having a specific capacitance in the range of 190–3480F/g with energy density ranging from 14–89 Wh/kg12. It has been reported that the capacitance due to faradaic peaks increases the specific capacitance of supercapacitor 10–100 times higher than the electrostatic capacitance of an EDLC. Recently, Ko et al.13. has demonstrated porous graphitic carbon with Ni2P2O7, heterostructure has shown high specific capacitance of 1893 F/g and Liao et al.14. has reported ultrahigh supercapacitance of 3480F/g using Co3O4 nanoparticles on vertically aligned graphene sheets supported in carbon fibers. Although pseudocapacitors are shown to have high initial capacitance, but due to limited durability, lack of stability during cycling and poor power density, its practical use in solid-state energy storage devices is limited16. In addition to this, crystallinity and morphology also play an important role in specific capacitance of pseudo capacitors, lower the crystallinity higher is the specific capacitance, which is due to the availability of chemically active dangling units which can easily take part in oxidation-reduction cycle8. To overcome these limitations, there is an extensive drive to develop EDLC type supercapacitor that carries various advantages over pseudo capacitor. In case of EDLC, electrical energy is stored over the layer of current collector without any redox couple. Due to the absence of faradaic current, EDLC type supercapacitors are more reliable and have shown long term durability with minimum loss of capacitance under multiple charging-discharging cycles.Current economy and population growth rates project a demand for energy of 40 terawatts (TW) with a total population of 10 billion by 2050et al. have fabricated graphene hydrogel for the EDLC type supercapacitor where storage performance is related to the high surface area17. Sundaram et al. have fabricated MnO2 nanostructures for EDLC type supercapacitance in a lower applied potential window having Csp of 50F/g18. Various carbonaceous materials till now have been developed for the testing of EDLC type supercapacitors due to its cost efficiency, high surface area, porous nature and versatile existing forms20. Significant enhancement in capacitive performance is not only governed by the properties of electroactive materials but also by the separators and electrolytes. Lin et al. have fabricated graphene based supercapacitor (Csp of 130F/g) in ionic liquid electrolyte21 where ionic liquids shows high chemical stability as compared to conventional electrolytes at a wide potential range22. Further development of the storage device lies on the various tricky parameters such as, proper choice of electrolyte, electrochemically stable current collector and thin porous spacer matrix between the electrode materials etc24. The current developmental trend of miniaturized autonomous electronic equipment such as implantable medical devices and active radio frequency identification (RFID) tags have raised the demand for solid-state-supercapacitors25. Significant efforts have been devoted to improve flexibility and energy storage/transfer capacity of solid-state supercapacitors based on various carbonaceous materials and their composites26, out of which graphene shows a promising choice of material due to high conductivity, flexibility, transparency, large surface area and layer like structure that drags attention to various scientific communities for its large scale application in energy storage devices24. In most of the cases, graphene based solid-state-supercapacitors demonstrate very low specific capacitance which is mainly due to the parallel restacking of graphene sheets resulting in decrease of the specific capacitance28. Recently, 3D graphene proves to be more prominent material for solid state supercapacitor due to the partial stacking of graphene sheets in random orientation in three dimensions resulting in formation of micro porous structures17. This unique orientation of graphene not only helps to improve the charge storage capacity of graphene but helps the electrolytic ions to move freely via porous geometry. To further improve the response of the 3D graphene as solid-state-supercapacitor various layer/porous nanomaterial hybrids have been reported which develops the charge storage capacity of the 3D graphene via increase in surface area and porous nature of the hybrid materials33. It is already reported that graphene based hybrid material possess superior charge storage capacity by providing enhanced surface area, electrical conductivity, thermal stability and mechanical strength to graphene34. Out of the various nanomaterials, 2D transition-metal dichalcogenides have been a perfect candidate for charge storage applications due to their layer like structures that shows weak Van der Waal force of attraction with graphite or graphene, especially 1T-MoS2 nanostructures41. Due to their band gap of ~1.9 eV, which reveals the semi-insulating nature, they are thus not immediately thought to be an electrode material for energy storage application42. But still MoS2 nanosheets display an excellent performance in wide, negative potential in neutral electrolytes, such as high specific capacitance and rate performance, which promotes the MoS2 nanosheets to be a promising electrode material for supercapacitor applications43. Till date various MoS2 based supercapacitor have been developed but only handful reports have been presented which shows the practical applicability of solid state supercapacitor46. The main research challenges associated with electrochemical supercapacitors are to develop new electrode materials, to enhance specific capacitance by modification of the electro- active material37.Xu 2 nanoflowers with 3D graphene heterostructure as an active material for energy storage device. The real demonstration of the solid state supercapacitor has been performed here using the 3D graphene-MoS2 hybrid over graphite current collector. Herein, the specific capacitance of bare MoS2 nanostructures comes out to be 169.37F/g with an energy density of 28.43 Wh/kg and power density of 10.17 W/kg. The capacitive performance of 3D graphene-graphite electrode has been improved significantly when the current collector is incorporated with MoS2 nanoflowers. This remarkable supercapacitor performance of the hybrid could be attributed to the combined effect between layered MoS2 and 3D graphene which could have some real phase application in modern energy storage devices.In this work, we report the simpler and cost effective process via chemical route of synthesis of hybrid MoS2 nanoflowers were characterized by XRD, SEM, and Raman spectroscopy. Figure 2 nanoflowers. It shows the diffraction patterns at 2θ = 13.55, 33.06,35.22, 42.5, 49.82, 58.01, and 69.77 which arises from the , (1 0 2), (006), (105), (110) and (201) planes of MoS247. The observed diffraction patterns are broadened in nature with a slightly lower diffraction angle shift as compared to the bulk 2H-MoS2, suggesting the presence of randomly stacked layers of MoS2 which may be due to the intercalation of NH3 in between the layers during the synthesis48. The XRD pattern well matches with the already common works on randomly stacked MoS2 nanostructures50. No peak other than MoS2 has been observed in the XRD pattern revealing that the synthesized MoS2 nanoflowers are highly pure in nature. Figure 2 nanoflowers at a low magnification, it is observed that the particles are spherical in nature with porous nanostructure in the order of nanometers. It is well established in the literature that porous nanomaterials possess a superior charge storage capacity as compared to that of their bulk counterpart51. The formation of the porous MoS2 nanoflowers is due to the reaction conditions as well as the precursors used in the reactions. Coagulation into a sphere is most likely due to the hydrothermal reaction conditions and the precursors used. During the reaction, ammonium molybdate releases MoO4−. These ions, having a layered structure, react with sulfide ions and intercalation by the residual ammonia prevents the stacking of MoS2 layers which leads to the self-assembly of a sphere-like morphology48. The Raman spectrum of MoS2 nanostructure was carried out at room temperature , arises due to S atom in the basal plane. Band appearing at 347.86 cm−1 (E2g symmetry) arises due to intralayer vibrational mode of Mo and S atoms in the basal plane. The A1g mode which appears at 390 cm−1 is due to the intralayer mode involving the motion of S atoms53. Figure −1, respectively56. The appearance of D band arises due to the activation of the first order scattering process of sp3 hybrid carbons, which is attributed to dislocation defects in graphene sheets57. Similarly, the SEM studies of as-synthesized 3D graphene were carried out, see  planes of MoS2 and multilayer graphene59.The MoSial Fig.  demonstr2 modified glassy carbon electrode in 0.1 M KOH solution from low (10 mV/s) to higher (300 mV/S) scan rate. From the CV it is clear that no significant redox peak was observed which reveals that MoS2 nanostructure behaves as typical electrical double layer capacitance which is similar to already published in the literature60. Also the working potential of MoS2 nanoflowers were quite high i.e. −0.9 to 0.2 i.e. 1.1 V of working potential was observed. The specific capacitance of MoS2 was calculated using given equation:61. By this equation, the specific capacitance of MoS2 nanostructure comes out to be 145.73F/g (100 mV/s).Figure 62:The energy density of the present system was calculated by using equation2 nanospheres in 0.1 M KOH solution at various scan rates. As seen from the figure the approximately rectangular shape with high symmetry is an indicator to double layer capacitance with fast charge-discharge rates. These results i.e. electrical double layer formation, are in good agreement with the previously published literature63. For various scan rates i.e. 100, 150, 200, 250, 300 mV/s, the specific capacitance using equation  in 0.1 M KOH. By comparing their respective cyclic voltammograms, it is observed that the hybrid material not only increases the working potential window but also increases the current response, Csp and energy density for graphite, 3D graphene over graphite and of 3D graphene-MoS2 hybrid comes out to be 7.83, 46.82, 58.0 F/g and 1.09, 13.0, 24.59 Wh/Kg, respectively which shows that the hyrid material has more supercapacitive performance as compared to both 3D graphene and graphite material. The complete values of specific capacitance, energy density, and power density was given in Tables The specific capacitance obtained by from CV curves is about 169.37, 95.66, 62.52 and 46.17F/g for 1, 2, 3 and 4 A/g respectively. This observed specific capacitance shows a significant improvement over previously reported valueseet Fig. . Green, 2 was scotch taped and then put in teflon line stainless steel vessel, 5 mg/mL solution of graphene oxide was added along with 2 M ascorbic acid solution and afterwards hydrothermal was sealed. The hyrothermal bomb was then kept at 120 °C for 4 h resulting in the formation of hydrogel of 3D graphene over graphite piece. This modified graphite was kept in 5 mg/mL solution of MoS2. Similiary other plate of supercapacitor was fabricated using same protocol. After this the two plate was assembled by using filter paper soaked with PVA/KOH solution.Figure 2 comes out to be 74.94F/g and the specific capacitance of 3D graphene-MoS2 hybrid in solid state device comes out to be 58.0F/g  fully opens the stacked 3 D graphene as well as MoS2 nano flaks which allow ion intercalation and creates abundant ion-accessible sites for adsorption/intercalation, and dramatically increases the cell’s capacitance68.Successful implementation of the supercapacitor in solid-state without a loss in performance can be inferred. Figure see Fig. . This is2 3D graphene hybrid over graphite. TGA spectrum shows two weight losses one at ~350 °C and other at around 500 °C. Weight loss i.e. 5% at 350 °C corresponds to 3D graphene69. While the weight loss i.e. 2.5% at around 500 °C corresponds to MoS2 nanoflowers59. From the TGA calculation, the weight of the active material comes out to be 1.1 mg, which has been used to calculate the specific capacitance  is performed by electroactive materials to analyze the kinetic feature of the ion transportation and ion diffusion in the porous hybrid electrode in aqueous electrolyte system. Figure s) for MoS2-3D graphene hybrid is 18.66 ohm. The corresponding equivalent circuit has been shown in Figure 70. The enchantment in energy density is found to be attributed to the enlarge operating potential voltage of 2.7 V in the gel electrolyte (PVA with KOH) and the relatively high specific capacitance of both electrodes, especially the negative electrode, was found which is very important for the solid state devices. The stability curve strongly indicates that about 90% of the initial capacitance can still be retained after 1400 cycle at the scan rate of 100 mV/s  in KOH-PVA medium without utilizing any expensive ionic liquid mediums. Recently a solid state architecture has been designed using a similar heterostructure66, however a costlier growth technique has been adopted where for the graphene growth was performed in 1000 °C in CVD setup. In contrast our study shows a very simpler and cost effective process via chemical route to design heterostructure. In comparison to the device performance, they report the specific capacitance of 19.44 F/cm3 in contrast to our device as 58.0 F/g. Here, it is noticed that the straight forward comparison is not feasible because the analytical approaches are entirely different.The Table 2 was successfully paired with 3D graphene to fabricate a flexible, solid-state supercapacitor with outstanding performance. The facile method of fabrication can easily be scaled up at minimal cost to create a large array of supercapacitors which is lightweight and reliable. An immediate application can be found in solar energy storage due to its large energy storage capacity in a short span of time in purely EDLC type nature. The specific capacitance of the solid state supercapacitor comes out to be 58 F/g with energy density of 24.59 Wh/Kg and power density of 8.8 W/kg. It can be a partial replacement of conventional batteries and can be easily incorporated into flexible and thin electronics due to its high stability, thinness and flexibility. The operating potential window can be further enhanced by using an ionic-liquid gel-based electrolyte. Other transition metal dichalcogenides can also be investigated along with MoS2 for their supercapacitor performance on top of 3D graphene matrix.MoSGraphite, ammonium molybdate, ascorbic acid, isopropyl alcohol, thiourea were purchased from sigma aldrich. All the solutions were prepared in deionized water. KOH was purchased from TCI chemicals.2 nanoflowers were synthesized by a hydrothermal method using ammonium molybdate and thiourea as a starting precursor. Briefly, 1 M solution of ammonium molybdate and 5 M solution of thiourea were prepared separately and transferred into a teflon-lined stainless steel autoclave (100 mL capacity) and hydrothermal reaction was carried out at 180 °C for 24 h. Black precipitate was obtained followed by washing with deionized water and absolute ethanol. The washing procedure is repeated several times until the final product is free of any trace amount of residual ions. The final product was dried at 80 °C for 12 h.The MoS2 nanospheres were coated on a polished GC electrode surface  to modify the Glassy Carbon (GC) electrode. For the coating, a solution of 1 mg/mL of MoS2 was prepared in isopropyl alcohol from which 5 µL was drop casted over the GC electrode surface. The prepared electrode was kept in a vacuum desiccator for 24 h to form a uniform layer over the surface. Cyclic voltammetry was carried out using a CHI electrochemical workstation with a three electrode system. While the MoS2-modified electrode was the working electrode, Ag/AgCl in sat. KCl was used as the reference electrode and platinum electrode was the counter electrode. These were dipped in 0.1 M KOH electrolyte.The as-synthesized MoS2 in isopropyl alcohol for 12 h. The graphite pieces were finally taken out and dried in an oven at 60 °C for 12 h.We followed the modified hydrothermal reduction method for the deposition of 3D graphene onto the graphite electrodes. Briefly, two rectangular pieces were taken from a graphite sheet and they were dipped into 10 mL of 5 mg/mL aqueous dispersion of graphene oxide (GO). This was freeze dried at −20 °C for 12 h. After the solution temperature was brought to room temp, 0.6 ml of 2 M ascorbic acid (AC) was added to the graphene oxide aqueous dispersion and the mixture was sealed in a teflon-lined autoclave and heated in an oven at 120 °C for 4 h, followed by drying of the attached graphite pieces in an oven set at 60 °C for 6 h. Once the drying was completed, the graphite pieces were dipped into a 5 mg/mL dispersion of MoS0.5 g of poly-vinyl alcohol (PVA) was added in small amounts to 9 mL of boiling de-ionized water under constant stirring with a magnetic heater-stirrer. Once the PVA gel reached a uniform consistency and bubbles started to form, 1 ml of 0.5 mg/mL aqueous solution of KOH was added into the gel drop wise under constant stirring. On attaining homogeneity, a piece of whatman filter paper was cut, soaked in the gel and placed in between the two electrode pieces. This arrangement was left to dry for 12 h and hence, the final device was completely fabricated.Supplementry videoSupplementry Information


Graphene has been intensively studied for applications to high-performance sensors, but the sensing characteristics of graphene devices have varied from case to case, and the sensing mechanism has not been satisfactorily determined thus far. In this review, we describe recent progress in engineering of the defects in graphene grown by a silica-assisted chemical vapor deposition technique and elucidate the effect of the defects upon the electrical response of graphene sensors. This review provides guidelines for engineering and/or passivating defects to improve sensor performance and reliability. Graphene, a two-dimensional (2D) zero-gap semiconductor, has drawn great interest as a promising platform for novel electronic, optoelectronic, and energy harvesting systems –9. In pa− ions to edge defect sites, but the binding characteristic was not thoroughly determined. Nevertheless, existing methods for introducing defects in graphene entail difficulties in controlling the quantity of the defects and/or avoiding contamination from external substances . I. I30]. I3 atmosphere. In the drain current versus gate voltage (Id–Vg) curves of graphene mesh FETs, the Dirac point was shifted greatly in the negative direction after N doping. During the annealing process, physisorbed molecules such as O species [Because the graphene meshes are directly synthesized, their edges are less vulnerable to contamination than those produced by lithographic methods. To further examine the characteristics of the edge defects, doping of graphene meshes was explored by high-temperature thermal annealing under NH species , 53 were2, even at room temperature [2 sensing. However, rigid Pd films undergo structural degradation during reaction with H2 [Pd shows high reactivity and resistance change upon exposure to Hperature –57, maki with H2 , 57–60.  with H2 , Pd-Si [ with H2 , and Pd- with H2 –65. In t with H2 , 67. Acc2 gas sensors have been fabricated with normal Pd-Gr and with Pd-GM [2 gas, and the continuous changes in resistance were monitored at room temperature. The relative resistance changes were greater for Pd-GM sensors upon exposure to H2 gas of concentrations ranging from 2 to 15 ppm /R0, where R0 and R respectively denote the channel resistances before and after exposure to H2 gas, improved by 18.2–28.8 % when graphene was replaced with graphene mesh  showed that Pd-GM sensors responded slightly faster than Pd-Gr to H2 gas  provide binding sites for the H+ ions, thereby further increasing the Dirac point shifts in the GM-FETs.To investigate the effect of defects in graphene-based FET pH sensors, electrolyte-gated graphene-FET (Gr-FET) and graphene mesh–FET (GM-FET) devices have been tested . Both se+ ion binding upon the cycling behaviors of graphene-based FET sensors. During cyclic tests in which GM-FET sensors were repeatedly exposed to conditions of various pH from 8.25 to 6.55, large Dirac point shifts of 90 mV were observed in the first cycle (Fig. d–Vg characteristics in response to pH became reversible, with corresponding sensitivities of ~7 mV/pH (Fig. Further analysis has been carried out to examine the influence of defect–H+ ions, whereas the increased number of defects at the mesh edges enhanced the pH response of the GM-FETs during the initial cycles. Furthermore, the irreversible response is believed to be caused by the direct adsorption of ions onto defects, which presumably involved strong chemisorption. It has been reported that H+ ions were attached so strongly to the edge defects and have only been detached after high temperature annealing [Gr-FETs have shown results consistent with those of GM-FETs: there was an irreversible portion of the response that gradually decreased with cycling, and only a reversible component remained after five cycles Fig. d–f. The nnealing . Such st+ ions at edge defects, which results in increased sensitivities that sometimes exceeded the Nernst limit. Unfortunately, such direct interactions involve irreversible covalent bonding, and therefore are not preferred in multiple-cycle sensor operations. For graphene mesh pH sensors, this issue has been solved through a simple process of passivating the edge sites. These results suggest how graphene mesh edge defects can improve sensors’ sensitivity and response time and also enable stable multiple-cycle operation through indirect carrier injection. As an example, biological sensors can be constructed by including graphene mesh for enhanced sensing characteristics, while improving stability by attaching appropriate receptors to the graphene edge sites to inhibit direct carrier injection.In this review, we investigated the defects of graphene meshes and their influence upon various sensor applications. Direct growth of graphene mesh by silica-assisted CVD is an excellent way to produce graphene with contamination-free defect sites. When these defects are introduced to Pd-GM gas sensors, the lowered energy barrier at the junctions between Pd nanoparticles and edge defects enhance indirect charge carrier injection into the graphene channel. As a result, the sensitivity and response time of Pd-GM gas sensors have been greatly improved over those of other graphene-based sensors. In contrast, graphene mesh pH sensors respond directly to H


This systematic analysis aims to assess the efficacy of PD-1/PD-L1 blockades compared with non-PD-1/PD-L1 therapy and investigate the potential predictive factors in epithelial carcinoma patients.p < 0.001 for OS and 0.66 , p < 0.001 for PFS. In subgroup analyses, HRs were 0.58 , p < 0.001 in PD-L1 ≥ 1% group, 0.75 , p = 0.001 in PD-L1 < 1% group for OS and 0.59 , p < 0.001 in PD-L1 ≥ 1% group, 0.80 , p = 0.136 in PD-L1 < 1% group for PFS. The p values of pooled HRs for OS in different age, sex and ECOG score groups were less than 0.001. In NSCLC patients, aggregated HRs for OS were 1.40 , p = 0.114 in EGFR mutant group and 0.88 , p = 0.536 in never smokers.A total of 11 trials with 6716 patients of melanoma, non-small cell lung cancer (NSCLC) and renal cell carcinoma (RCC) were included. The pooled HRs (95%CI) were 0.67 , A systematic search from January 2010 to April 2016 was conducted for eligible clinical trials. Based on the data of hazard ratios (HRs) and 95% confidence intervals (CIs) for overall survival (OS) and progression-free survival (PFS), we assessed the pooled HRs and proposed the subgroup analyses.EGFR wild type were associated with extended OS in NSCLC patients.PD-1/PD-L1 blockades prolonged OS and PFS in epithelial carcinoma patients. PD-L1 expression was a predictive factor for PFS but not predictive for OS. Age, sex and ECOG score were excluded to predict any of the efficacy endpoints. Smoking history and  The checkpoint immunotherapy has been increasingly understood and used to unleash the immune system to fight against cancer . These yAs one of the most critical checkpoint immunologic treatments, PD-1/PD-L1 blockade has become a promising focus of immunotherapy in cancer treatment . Two antAlmost each of PD-1/PD-L1 blockades has satisfying overall response rates in treating different types of epithelial carcinoma , 7. HoweEGFR mutation (within NSCLC patients) and BRAF mutation (within melanoma patients) are also conducted to provide further evidence for clinical treatment.In consideration of predictive factors, we analyze different membranous PD-L1 expression levels, because PD-L1 expression of tumor cells is most abundant in epithelial carcinoma and most closely correlated with response to anti-PD-1/PD-L1 agents , 8, 9. BCochrane handbook for systematic reviews of interventions. The study selection process was presented in Figure According to the outlined search strategy, a total of 820 records were obtained, of which 371 duplicates were removed. After screening, 484 articles including reviews, case reports and non randomized controlled trials were excluded. Of the rest 19 records, 8 studies did not report the relevant data. Upon the remaining 11 studies, the two reviewers had the perfect agreement on their eligibility and assessed the quality of included studies independently by the scoring criteria stated in n = 3510), 4 in NSCLC patients (n = 2385), and 1 in RCC patients (n = 821). 3 of the trials were phase 2 trials, 1 was phase 2/3 trial, and 7 were phase 3 trials.The analyses were based on data from a total of 6716 patients enrolled in 11 randomized controlled trials. The experimental treatment drugs of those trials were PD-1/PD-L1 blockades, including nivolumab, pembrolizumab and atezolizumab, while the controlled interventions were standard chemotherapy , targeted therapeutic agents (everolimus) and other form of immunotherapy (ipilimumab). According to currently completed trials focused on epithelial carcinoma, all randomized controlled trials were conducted within melanoma, NSCLC and RCC patients. 6 of the enrolled trials were in melanoma patients  for OS and PFS of patients. For the PD-L1 expression evaluation, the immunohistochemistry assays of PD-L1 employed in the selected studies contained Dako, clone 28-8 (Epitomic) and 22C3 antibody (Merck). We retrieved the corresponding HR estimates with the cut-off of 1%, which meant membranous PD-L1 staining in at least 1% of tumor cells. The information of included studies' authors, cancer types, numbers of patients, interventions, basic characteristics of patients, and HRs for OS and PFS of PD-1/PD-L1 treatment p < 0.001) without significant heterogeneity (I2 < 0.1%), which reflected that compared to non-PD-1/PD-L1 therapy, PD-1/PD-L1 blockades reduced 33% in risk of death among epithelial carcinoma patients. This benefit had met the criteria of treatment superiority.The data available on OS pooling were from 10 observations. The pooled HR for OS Table  was 0.67p < 0.001). However, a considerable heterogeneity with I2 = 79.7 % was observed with the random effect model  for PFS in melanoma, NSCLC and RCC were 0.54 , 0.84 , 0.84  respectively.All studies reported the data on PFS, and the combined HR for PFS with 15 records was 0.66,  in PD-L1 ≥ 1% group, 0.75,  in PD-L1 < 1% group for OS and 0.59,  in PD-L1 ≥ 1% group, 0.80,  in PD-L1 < 1% group for PFS with not important heterogeneity within all subgroups .To investigate predictive role of PD-L1 expression, we computed the aggregated HRs for OS and PFS in different levels of PD-L1 expression  groups and generated the forest plot Figure . Pooled We also analyzed the baseline data such as age, sex and ECOG score of 0 or 1  to further explore other factors. The results of pooled HRs for OS and PFS corresponding to these factors were listed in Table p < 0.001) and non-squamous patients ; p < 0.001) had more extended OS when compared with non-PD-1/PD-L1 therapy. In EGFR wild type patients, the HR for OS was 0.66, ; p < 0.001, indicating the better efficacy of PD-1/PD-L1 blockades in those patients. However, in patients with EGFR mutant type, the HR for OS was 1.40,  and p was 0.114 (I2of heterogeneity = 67.3%), suggesting that anti-PD-1/PD-L1 therapy functioned not distinctly better than the control group treatment. Similarly, the never-smokers were verified not to have the expectedly longer OS with HR pooled as 0.88, ; p = 0.536 with I2 = 39.0%. The HR for OS in smokers was 0.71, ; p < 0.001 with the meaning of better outcomes due to the use of anti-PD-1/PD-L1 agents. Besides, in melanoma patients, both the BRAF wild type and BRAF mutant patients benefited in OS, and HRs (95%CIs) were 0.51, , p < 0.001; 0.55, , p < 0.001 respectively with not important heterogeneity   = 0.67, ; %CI = 0.6All reported data of the meta-analyses results were listed in Table The sensitivity analyses were conducted by excluding studies one by one. The results of those analyses showed no significant differences when compared to the former summary estimates and had excellent stability.p < 0.05) in our main analyses. Nevertheless, two possible biases with p < 0.05 by Egger's test were observed in the subgroups of Age < 65 and Male patients in the pooling process of HR for PFS.We assessed the publication bias of included data for pooled analyses using the Egger's test, and the p values were listed in Table It was acknowledged that PD-1/PD-L1 blockade, as a type of immune checkpoint inhibitors, had remarkable response rate and clinical results in patients with different kinds of cancer, especially those with epithelial-originated malignancies. Nonetheless, it remained unclear whether anti-PD-1/PD-L1 therapy, in contrast with other therapy, functioned better to extend OS and PFS and which subgroups of patients would benefit from the treatment. Our meta-analysis integrating all data from relevant trials were requested to solve the problem. To our knowledge, the meta-analysis was the first study to investigate the outcome and predictive biomarkers for PD-1/PD-L1 therapy in epithelial carcinoma patients within solely randomized controlled trials by assessing the two primary endpoints of OS and PFS.Our results provided convincing evidence that the OS of patients given PD-1/PD-L1 inhibitors was significantly longer than patients given other drugs as those individual studies reported (only one study comparing nivolumab with ipilimumab (CTLA-4 antibody)  excludedIn PD-L1 expression investigation, we found it not practicable to use PD-L1 as a biomarker to predict OS benefit when comparing PD-1/PD-L1 blockades with other control therapy, for both the higher (PD-L1 expression ≥ 1%) and lower (PD-L1 expression < 1%) expression groups could gain obvious clinical benefit from PD-1/PD-L1 blockades. Be that as it may, the higher expression group had the better PFS outcome, but the lower expression group was associated with insubstantial improvement of PFS. The immunohistochemistry (IHC) cut-off value (ranging from 1% to 50%) we chose to define the PD-L1 positivity was 1%, in that the clinical data assessed by this point were the most abundant in the included 11 trials. Besides, cutting off by the lowest expression level enabled to include the most possible patients who could benefit from the therapy. However, the assessment of PD-L1 expression was much complicated because the tumor PD-L1 expression was not constant which was associated with activated tumor antigen-specific T cells  and coulBRAF mutant or BRAF wild type patients in melanoma group, and the squamous cancer patient or non-squamous cancer patients in NSCLC group had gained better outcome of survival. But there was no significant improvement for OS in patients without smoking history and patients with EGFR mutations. Whilst, the present or previous smoker, patients with EGFR wild type showed longer survival time, implicating the smoking history and EGFR wild type might be considered as the potential predictive factors for anti-PD-1/PD-L1 treatment. In our speculation, the association of mutations and other exposures to mutagens like smoking with the efficacy of PD-L1 blockades was possibly because the tumor antigen, considered as the target of T cell activated by checkpoint blockade, was related to the consequence of somatic mutations [The next novel finding of our analyses was in different subgroups. We found that the utations , 21, 22.Finally, OS had been improved in the overall patient regardless of the age, sex and ECOG score. Thus, it was persuasive that those factors were not meaningful indicators for the eligibility of anti-PD-1/PD-L1 treatment.EGFR wild type were potential indicators for prolonged OS in NSCLC patients. There were multiple clinical trials ongoing and many other antibodies targeting PD-1/PD-L1 under early-stage development currently, more comprehensive data from future clinical trials focused on this field were still needed for further investigation.In conclusion, the aggregated HRs for OS and PFS summarized in our systematic analyses revealed that in the comparison of anti-PD-1/PD-L1 agents with other control therapy, the PD-L1 expression was not an appropriate factor to predict the benefit of OS in epithelial carcinoma patients, but could be predictive for PFS. Age, sex and ECOG score were excluded to predict any of the outcome endpoints. Smoking history and We searched for the articles of clinical trials from PubMed, Embase, Web of Science, and the Cochrane Library from January 2010 to April 2016. Records at American Society of Clinical Oncology (ASCO), the European Society for Medical Oncology (ESMO), the world Conference of Lung cancer (WCLC) were also reviewed. The following search terms were used: “pembrolizumab”, “Nivolumab” “atezolizumab”, “Tremelimumab”, “AMP-224”, “MDX-1105”, “pidilizumab”, and “cancer/carcinoma”.All relevant articles underwent evaluation for eligibility by two investigators independently and we selected the articles according to the following criteria: 1) articles with randomized controlled trials (RCTs); 2) at least one of the two endpoints  reported; 3) published in English; 4) the full text available. Our exclusion criteria were as below: 1) letters, expert opinions, case reports and reviews; 2) articles without available data; 3) duplicate publications.Cochrane Handbook for Systematic Reviews of Interventions , and evaluated the random sequence generation, allocation concealment, blinding of participants and personnel, blinding of outcome assessment, incomplete outcome data, selective reporting and other bias to ensure the low-risk of bias of the studies included.We assessed the quality of involved randomized controlled clinical trials according to the criteria presented in the The data of study identification, the intervention of experimental treatment and control group, numbers of enrolled patients in each trial, patients' detailed information , hazard ratios(HR) with their 95%CIs and p values for OS and PFS were extracted by two individual investigators independently. We also collected the relevant information in every subgroup we set to render sufficient data to our subgroup analyses.2 statistic were used to indicate the heterogeneity. A p value less than 0.05 in the Q test or an I2 value greater than 50% in the I2 statistics suggested the significant heterogeneity. If the heterogeneity was significant, we used the random effect model of pooling instead of fixed effect model and designed subgroups analyses to clarify the between-study heterogeneity. To test the publication bias of the included studies, the Egger's test was chosen. All the statistical analyses were performed with STATA/SE software version12.0 .We calculated pooled HRs and their 95%CIs for OS and PFS which were considered to be the primary outcome of the meta-analyses and generated the forest plots accordingly. The chi-square Q test and I


SQSTM1) and valosin-containing protein (VCP) are 2 key genes associated with several neurodegenerative disorders but have yet to be thoroughly investigated in sIBM. A candidate gene analysis was conducted using whole-exome sequencing data from 181 sIBM patients, and whole-transcriptome expression analysis was performed in patients with genetic variants of interest. We identified 6 rare missense variants in the SQSTM1 and VCP in 7 sIBM patients (4.0%). Two variants, the SQSTM1 p.G194R and the VCP p.R159C, were significantly overrepresented in this sIBM cohort compared with controls. Five of these variants had been previously reported in patients with degenerative diseases. The messenger RNA levels of major histocompatibility complex genes were upregulated, this elevation being more pronounced in SQSTM1 patient group. We report for the first time potentially pathogenic SQSTM1 variants and expand the spectrum of VCP variants in sIBM. These data suggest that defects in neurodegenerative pathways may confer genetic susceptibility to sIBM and reinforce the mechanistic overlap in these neurodegenerative disorders.Genetic factors have been suggested to be involved in the pathogenesis of sporadic inclusion body myositis (sIBM). Sequestosome 1 ( Muscle SQSTM1 had not been investigated in sIBM until a recent study using targeted next-generation sequencing in a group of 79 patients  (VCP) gene are known to cause an inherited form of IBM with Paget disease and frontotemporal dementia (IBMPFD) , has been recognized as a strong biomarker in muscle with a high sensitivity and specificity for sIBM . Geneticpatients . In thatmyopathy . In addiia (FTD) . Mutatio(IBMPFD)  and have(IBMPFD) . Two misdementia .SQSTM1 and VCP genes in sIBM, we investigated these 2 genes using whole-exome sequencing data from 181 sIBM patients, which was produced as a part of an International IBM Genetics Consortium.These findings, along with denervation in muscle electromyography of sIBM patients, suggest a possible genetic overlap between sporadic and IBM-like myopathies and also neurodegenerative diseases. To thoroughly investigate the contribution of 22.1N = 235) aged >60 years were used as an internal aged control group to compare with our sIBM cohort. The study was approved by the National Research Ethics Service Committee London—Queen Square (research ethics committee reference: 12/LO/1557).This study is part of the International IBM Genetics Consortium, a Muscle Study Group–endorsed project, which currently has members from 17 specialized centers in 7 countries around the world. For this whole-exome sequencing Consortium study, DNA samples from a total number of 181 sIBM patients were collected from 11 centers. Patients diagnosed with sIBM had to have an sIBM diagnosis according to a muscle diseases expert and also had to fulfil the Griggs criteria , the Eur2.2SQSTM1 and VCP genes. We excluded all synonymous variants, and all common variants with a population frequency >1% were identified in the 1000 Genomes project (www.1000genomes.org/), in the Exome Variant Server (EVS) database (evs.gs.washington.edu/EVS/), in the Exome Aggregation Consortium (ExAC) Browser (exac.broadinstitute.org/), and in the internal aged controls, as these variants less likely play a role in a rare disease. The filtered variants were confirmed by the conventional Sanger sequencing. The allele frequency of each variant found in sIBM was compared with the ExAC database using Fisher test. The pathogenicity of these variants was evaluated using the following in silico prediction tools: SIFT , and the concentration, purity, and integrity of each RNA sample were assessed as previously described , 2 patients with VCP variants (cases 6 and 7), and 5 age- and gender-matched controls. Raw expression data were log2 transformed and quantile normalized, and differential expression analysis  was performed using the limma Bioconductor package  pathways, and Human Phenotype Ontology terms was performed using g:Profiler (biit.cs.ut.ee/gprofiler/). Among the upregulated genes, 3 genes associated with inflammation markers  were selected for validation using real-time quantitative polymerase chain reaction (RT-qPCR). Briefly, total RNA (600 ng) from the 6 cases and 8 controls was reverse transcribed into complementary DNA using random primers from High-Capacity cDNA Reverse Transcription Kit . Three replicates per sample were assayed for each target gene using Fast SYBR Green PCR Kit (Applied Biosystems) and run in a QuantStudio 6 Flex Real-Time PCR System (Applied Biosystems). Details on primers and RT-qPCR conditions are available on request. Cyclophilin (PPIA) was selected as the reference gene. The 2−ΔΔCt method .Whole-genome expression profiling was performed using the Illumina HumanHT-12 v4 Expression BeadChip  on 3 patients with  package . Genes wt method  was used3From the entire cohort of 181 sIBM patients, 150 (82.9%) were Caucasians and 16 (8.8%) were from other ethnicities, including Asian Chinese, the Indian subcontinent, and Black Africans (ethnicity information was unavailable for 15 patients). The majority of sIBM patients were male (65.7%). Age of onset, which was collected retrospectively, ranged from 31 to 85 years (mean 59.6 ± 9.6 years). The mean age of the 235 healthy aged controls was 79.1 ± 8.5 years, ranging from 60 to 102 years, and similarly to the sIBM cohort, the majority were also male (61.7%).SQSTM1 gene (ENST00000389805) were found in 4 patients (VCP gene (ENST00000358901) were found in 3 sIBM patients. Of note, the frequency of the variants SQSTM1 p.G194R and VCP p.R159C was significantly higher in our sIBM cohort compared with the ExAC database . These 2 variants were absent in the other population databases and in our aged control group. From the 6 rare variant we found, 4  from patients with VCP variants prevented statistical analysis of this sIBM patient group. The expression of SQSTM1 and VCP did not show significant differences between any patient group and controls, but a significant upregulation of MHC genes (class I [HLA-A] and class II [CD74 and HLA-DRA]) was seen in the group of patients carrying SQSTM1 variants , which is a multifunction protein participating in a number of different biological pathways  and II (HLA-DRA) genes were significantly upregulated in patients compared with controls by RT-qPCR, which is consistent with a previous study (CD74) was significantly upregulated only in the SQSTM1 patient group. The statistical analysis could not be carried out for the VCP patient group because of small sample size, but there was also a trend for the upregulation of these inflammation markers. The more pronounced upregulation observed in the SQSTM1 sIBM patient group than in the VCP patient group. This is the first time that different expression level of inflammation markers between patients with sIBM is suggested. Although further cases should be analyzed, the MHC expression could be a potential differentiating factor that directs the clinical phenotype of SQSTM1 or VCP toward sIBM as opposed to the other neurodegenerative conditions. Additional analysis in muscle and other tissues of patients carrying SQSTM1 or VCP mutations without symptomatic sIBM but other phenotypes is also necessary in the future to confirm this hypothesis.The MHC class I (us study , and theSQSTM1 variants and expand the spectrum of VCP variants in sIBM. Our findings suggest that variants in these genes constitute genetic susceptibility factors for sIBM and for other multisystem proteinopathy phenotypes. The findings from this study also expand the clinicopathologic spectrum of diseases associated with SQSTM1 and VCP genes, and the overlap between sIBM and IBMPFD, ALS, and/or FTD suggests that muscle and brain diseases share similar pathogenic pathways that may be important for further biomarkers, genes, and therapeutic target discovery. Further investigation of the sIBM whole-exome sequencing data is still ongoing and data from this international collaboration will likely reveal further findings.In conclusion, we report for the first time likely pathogenic Declaration of interests: All authors have no competing financial interests. Authors' contributions: Qiang Gang contributed to sample collection, all the experimental work, data analysis, and drafting the first version of the manuscript; Conceição Bettencourt contributed to the experimental plans of microarray and RT-qPCR, preliminary analysis of microarray data, and statistical plan; Pedro M. Machado contributed to overall coordination of the study, namely worldwide sample collection and liaising with all the study collaborators; Conceição Bettencourt, Pedro M. Machado, and Henry Houlden also contributed to drafting the first version of the manuscript; Janice L. Holton contributed to the review and photography of the muscle biopsies; Alan M. Pittman and Deborah Hughes contributed to the generation of whole-exome sequencing data; Stefen Brady, Janice L. Holton, and Boel De Paepe contributed to immunotyping of muscle biopsies; and Andrew B. Singleton contributed to the exome data from control individuals. All the authors from International IBM Genetic Consortium and Muscle Study Group contributed to acquisition of clinical data and sample collection. Michael G. Hanna, Henry Houlden, and Pedro M. Machado are principal investigators of the International IBM Consortium Genetics Study. All the authors contributed to the critical revision of the manuscript and approved the final version.


Radiation therapy is critical for the control of many tumors and lung is an important dose-limiting organ that impacts radiation dose prescribed to avoid irreversible pulmonary fibrosis in cancer survivors. Idiopathic pulmonary fibrosis (IPF) is a chronic, irreversible lung disease caused by aberrantly activated lung (myo)fibroblasts. The presence of pro-fibrotic, apoptosis-resistant fibroblasts in IPF promotes progressive fibrosis and may have a role in other diseases, if these resistant cells are selected for as a consequence of treatment. However, the pathological response of IPF fibroblasts to radiation compared to non-IPF lung fibroblasts is not known. To address this, we examined fibroblast viability following radiation in lung fibroblasts from IPF and non-IPF patients and the underlying mechanism that protects IPF fibroblasts from radiation-induced death. IPF fibroblasts are significantly more resistant to apoptosis compared to non-IPF lung fibroblasts, suggesting that resistance to radiation-induced cell death is a predominant mechanism leading to lung fibrosis. Analysis of γH2AX induction demonstrated that radiation-induced DNA damage is reduced in IPF fibroblasts and correlates to the activation of the transcription factor forkhead box M1 (FoxM1) and subsequent upregulation of DNA repair proteins RAD51 and BRCA2. FoxM1 activation occurs secondary to FoxO3a suppression in IPF fibroblasts while restoration of FoxO3a function sensitizes IPF fibroblasts to radiation-induced cell death and downregulates FoxM1, RAD51, and BRCA2. Our findings support that increased FoxO3a/FoxM1-dependent DNA repair may be integral to the preservation of death-resistant fibrotic fibroblasts after radiation and that selective targeting of radioresistant fibroblasts may mitigate fibrosis. Rates of pulmonary fibrosis can be as high as 70–80% in high dose regions of irradiated lung3. It is currently unclear whether radiation-induced lung fibrosis (RILF) results from failure of normal healing after pneumonitis or is a separate, complicating event5. Thus, it is difficult to predict the true risk of RILF, for which there are no effective treatment strategies7. Recent work in idiopathic pulmonary fibrosis (IPF), a progressive and lethal lung disease, showed that fibroblasts derived from IPF patients maintain an apoptosis-resistant phenotype in response to various stressors14. Elucidation of this mechanism is crucial in understanding fibrotic disease, regardless of the inciting cause.Radiation therapy is prescribed in over 50% of patients receiving cancer treatment. Radiation-induced toxicities are relatively common following radiation when normal lung is in close proximity to tumor. While pneumonitis is an early and potentially reversible toxicity, pulmonary fibrosis is delayed, progressive and can impair normal lung function15. Fibroblasts become activated and produce collagen-rich extracellular matrix during repair of basement membranes15. Contrary to normal healing, thoracic radiation inappropriately activates myofibroblasts, which promote the deposition of type I collagen that destroys parenchyma and establishes a niche for ongoing fibrosis17. To further compound this injury, alveolar epithelial cells may undergo trans-differentiation into myofibroblasts in IPF and RILF18. The most lethal event following radiation to non-hematopoietic cells is the induction of DNA double-strand breaks (DSB), which induces mitotic catastrophe and apoptosis after 2–6 days23. Under normal physiologic conditions, DNA DSB trigger a cascade of events that encourage repair at the site of DNA damage24. Homologous recombination (HR) repair following DNA DSB is a primary, high-fidelity mechanism of radiation repair in human cells. An important step in HR is recruitment of the repair protein RAD51 by breast cancer-associated gene 2 (BRCA2) to the damaged DNA sites; the alteration of these proteins renders cells resistant to cytotoxic damage26. FoxM1, a member of the Forkhead family of transcription factors, is known to upregulate DNA repair proteins such as RAD51 and BRCA2, thereby protecting cells from radiation-induced DNA damage28. FoxM1 was increased in irradiated murine lung tissue and in human IPF fibrotic lesions; moreover, the conditional deletion of FoxM1 prevented lung fibrosis29. FoxM1 activation occurs following suppression of FoxO3a, indicating a negative feedback loop exists between these two family members30, and FoxO3a is aberrantly suppressed in IPF fibroblasts and patient IPF lung tissues32. We therefore sought to examine the FoxO3a/FoxM1-dependent pathway in IPF cells in response to ionizing radiation. We found that IPF fibroblasts sustain less radiation-induced DNA damage compared to non-IPF lung fibroblasts and are highly viable after radiation. We further demonstrated that enhanced FoxM1 in IPF fibroblasts leads to increased RAD51 and BRCA2 gene expression. Following FoxO3a over-expression or FoxM1 silencing, RAD51 and BRCA2 expression is abrogated and sensitizes IPF fibroblasts to radiation-induced death. We propose that FoxM1 may be pivotal in selecting apoptosis-resistant fibrotic fibroblasts within irradiated lung tissue, contributing to the development and progression of RILF. FoxM1 may therefore represent a therapeutic target as it regulates fibroblast survival following radiation damage.Ionizing radiation initially injures pulmonary epithelial cells, releasing pro-inflammatory cytokines that recruit inflammatory cellsAs radiation is a risk factor for organ fibrosis, irradiated fibrotic fibroblasts likely also acquire pathological properties and become resistant to death. To test this, radiation dose-response viability was measured in IPF and control lung fibroblasts in the presence or absence of collagen matrix. There was no significant difference between IPF and control fibroblasts when cells were cultured on a tissue culture plate Fig. . However34. We measured γH2AX as an indirect indicator of DNA DSB, which correlates to radiation-induced cell death36. Radiation upregulated γH2AX expression in both IPF and control fibroblasts at 1 h  molecules form around the break site to encourage accumulation of DNA repair proteins1 h Fig. , left, h1 h Fig. , right. 1 h Fig. . To furt1 h Fig. , we trea1 h Fig. , left. E1 h Fig. , right. 29. FoxM1 regulates the expression of several DNA repair proteins including RAD51 and BRCA2, which are important for proper DNA DSB repair28. We therefore examined the role of FoxM1 on the expression of RAD51 and BRCA2 in irradiated IPF fibroblasts. Increased FoxM1 levels were found in IPF fibroblasts between 1 and 6 h compared to FoxM1 levels of irradiated control fibroblasts . The tissue samples were stripped of all identifiers and designated as waste (exemption 4). Written informed consent was obtained on all patients prior to the procedure being performed. Use of human lung tissues was approved by the Institutional Review Board (IRB) at the University of Minnesota. The diagnosis of IPF was supported by history, physical examination, pulmonary function tests, and typical high-resolution chest computed tomographic findings of IPF. In all cases, the diagnosis of IPF was confirmed by microscopic analysis of lung tissue that demonstrated the characteristic morphological findings of interstitial pneumonia52. Removed lung tissues were chopped to 5 mm3 and cultured in Dulbecco’s modified Eagle’s medium  supplemented with 20% fetal calf serum  and 2% antibiotics for 4–5 weeks at 37 °C in a 5% CO2 humidified incubator to prepare IPF and non-IPF (control) lung fibroblasts. They were used at passage numbers 3–9 for all experiments.Lung tissues removed at the time of transplantation or death from non-IPF and IPF patients , 10% 10× DMEM (MilliporeSigma) and 10% 1×DMEM, and the pH was adjusted to 7.2 with 0.1 M NaOH. Cell culture dishes or 96-well plates were coated with collagen and incubated for a minimum of 2 h at 37 °C prior to use. Following 24 h, serum-free DMEM (SF DMEM) was used for experiments. Human Bronchial Epithelial Cell3-KT (HBEC3-KT) cell line was obtained from American Type Culture Collection  and cultured in airway epithelial cell basal medium (ATCC) supplemented with 1% antibiotics and bronchial epithelial cell growth kit (ATCC) at 37 °C.Radiation was delivered using 6 MV photons from a clinical linear accelerator  routinely calibrated and maintained to clinical standards. Cells were treated with increasing radiation doses (0–15 Gy). Culture plates were placed on 1.5 cm of tissue equivalent material and the gantry rotated to 180 degrees to ensure electronic equilibrium at the level of cells. Radiation was prescribed using a single source-to-surface distance technique.n = 8) and IPF fibroblasts (n = 8) were cultured on collagen-coated or non-collagen-coated 96-well plates in SF DMEM prior to radiation or bleomycin . Cells were then incubated for 3 h with 20 μL CellTiter Blue reagent  to assess viability or with 20 μL CellTiter 96 Aqueous One Solution (Promega) to assess proliferation for 3 h. Cell viability and proliferation were measured at 560 nm (excitation)/590 nm (emission) of fluorescence and at 490 nm of absorbance using a 96-well plate reader , respectively.Randomly selected non-IPF control lung fibroblasts (Deoxynucleotidyltransferase (TdT)-mediated dUTP nick-end labeling (TUNEL) assay was conducted using In Situ Cell Death Detection kit  according to manufacturer’s instruction. Briefly, 4 of each control and IPF fibroblasts plated on collagen matrix coated cover slips  in SF DMEM were irradiated with 15 Gy and cells were then incubated for an additional 24 h. Control cells showing low cell viability and IPF cells showing high cell viability in response to the radiation were chosen for the current assay. After the incubation, cells on cover slips were washed with phosphate buffered saline  and fixed with 4% paraformaldehyde solution  at 4 °C over night. Cells on cover slips were then permeabilized with Proteinase K  at room temperature for 20 min. After the washing with PBS, cells were incubated with TUNEL reaction mixture at 37 °C for 2 h and washed with PBS twice. In addition, the fixed control cells on cover slips was incubated with Nuclease  at 37 °C for 25 min before incubating in TUNEL reaction mixture to be used as a positive control. For fluorescence microscopic analysis, the cover slips were washed with PBS and sequentially moved on slide glasses having mounting solution with DAPI . The images were taken from two different areas on each cover slip using a confocal fluorescence microscopy  at ×200 magnification, and the images were processed using Progres Capture Pro V2.8.8 software . Images analysis was conducted using ImageJ program .n = 8, each) used in cell viability assay (6 × 105 cells/60 mm cell culture dish) were irradiated with  9 Gy and incubated for the assigned time periods. After the incubation, cells were lysed with 1×cell lysis buffer  containing protease inhibitor (Roche Applied Science) and phosphatase inhibitor . Cell lysates were collected, sonicated on ice for 15 s, and then denatured using 5×Laemmli buffer at 95℃ for 10 min. The lysates were separated by a gradient polyacrylamide gels having 5–20% polyacrylamide concentration and proteins on the gel were electrically transferred to a PVDF membrane  using a Protean III tank transfer system (Bio-Rad). After blocking step using 5% skim milk in TTBS  for 1 h at room temperature, the membrane was incubated with γH2AX , H2AX , FoxM1 , FoxO3a , RAD51 , BRCA2 , GAPDH , Lamin A/C  or β-actin  antibody diluted in TTBS containing 5% bovine serum albumin (BSA) at 4℃ for 24–48 h. After washing with TTBS three times, membranes were incubated with anti-rabbit or mouse IgG antibody conjugated with horseradish peroxidase (MilliporeSigma) diluted in TTBS containing 5% skim milk for 1 h at room temperature. The protein bands on a membrane were detected by ECL solution (ThermoFisher Scientific) using Chemi Doc-IT2 image analyzer , and quantified using VisionWorks LS program (UVP BioImage systems).Same control and IPF fibroblasts (n = 8) and IPF (n = 8) fibroblasts using NE-PER Nuclear and Cytoplasmic Extraction Reagents kit (ThermoFisher Scientific) according to the manufacturer’s instructions. Briefly, control and IPF fibroblasts (1.6 × 106 cells/10 cm cell culture dish) cultured on collagen in SF DMEM were incubated for 6 h after 9 Gy radiation. Cells were lysed with Cytoplasmic Extraction Reagent I and II. Cytosolic fraction was collected by centrifugation at 16,000×g for 5 min and the remaining pellet was lysed with Nuclear Extraction Reagent. Nuclear fraction was then isolated by  centrifugation at 16,000×g for 10 min. Both cytosolic and nuclear fractions were denatured with 5×Laemmli buffer at 95 °C for 10 min, and FoxM1 and FoxO3a protein expression was measured using Western analysis. Lamin A/C and GAPDH were used as a nuclear and cytosolic protein marker, respectively.Nuclear protein fractions were isolated from control  cultured (3 × 105 cells/35 mm cell culture dish) on collagen was extracted at specified time points following 9 Gy radiation using TRIGent . One µg of total RNA was synthesized into complementary DNA, and PCR was conducted using Step One Plus real-time PCR system  and Power SYBR Green PCR master mix (Applied Biosystems) under the following conditions: initial denaturation at 95 °C for 10 min then amplification by cycling 45 times at 95 °C for 15 s, 55 °C for 30 s, then 68 °C for 45 s. GAPDH expression was examined to normalize the copy numbers of each gene. Primer Sequences for PCR: FoxO3a , FoxM1 , RAD51 , BRCA2  and GAPDH .Total RNA from control or IPF fibroblasts (n = 4) were transfected with FoxM1, BRCA2, RAD51, or control siRNA  using Lipofectamin RNAiMAX (Life technologies). For overexpression of FoxM1, control fibroblasts (n = 4) were transfected with 2 µg empty pCMV3 or FoxM1-pCMV3 vector  using Lipofectamin 3000 (Life technologies) in Opti-MEM . For FoxO3a overexpression, IPF fibroblasts (n = 4) were infected with either 2 × 106 PFU of ad-GFP-FOXO3a  or Ad-GFP  according to the manufacturer’s instructions. For co-overexpression experiments, IPF fibroblasts (n = 4) infected with ad-GFP-FOXO3a were transfected with FoxM1-pCMV3 vector as described above. Cells were then cultured on collagen in SF DMEM for 24 h followed by irradiation (9 Gy). FoxM1 overexpression or silencing by siRNA was verified by measuring FoxM1 mRNA  cultured on collagen in SF DMEM were irradiated at 9 Gy followed by incubation for an additional 6 h at 37 °C. Cells were then fixed with 4.0% paraformaldehyde in PBS and permeabilized in 0.1% Triton-X (MilliporeSigma) in PBS for 15 min. The cells were incubated with the anti-RAD51 antibody  diluted 1:300 in PBS at 4 °C for 24 h. Cells were then incubated with an anti-rabbit Alexa Fluor 488 conjugated secondary antibody  diluted 1:250 in PBS for 20 min. Cells on collagen matrix were cover slipped with ProLong Gold Antifade Mountant with DAPI (ThermoFisher Scientific). From each sample, 3 of ×20 magnified images were analyzed using an upright microscope . Images analysis was conducted using ImageJ (NIH).Control and IPF fibroblasts , showing lowest, lower quartile, median, upper quartile and highest expression levels. Two-dimensional column graphs were prepared with Microsoft Excel. Comparison between unirradiated and irradiated fibroblasts or comparison between irradiated IPF and control fibroblasts as a function of time was conducted using two-tailed Student’s Supplementary Figure 1Supplementary Figure 2Supplementary Figure 3Supplementary Figure 4Supplementary Figure 5Supplementary figure legends


Graphene field effect transistors are becoming an integral part of advanced devices. Hence, the advanced strategies for both characterization and tuning of graphene properties are required. Here we show that the X-ray irradiation at the zero applied gate voltage causes very strong negative doping of graphene, which is explained by X-ray radiation induced charging of defects in the gate dielectric. The induced charge can be neutralized and compensated if the graphene device is irradiated by X-rays at a negative gate voltage. Here the charge neutrality point shifts back to zero voltage. The observed phenomenon has strong implications for interpretation of X-ray based measurements of graphene devices as it renders them to significantly altered state. Our results also form a basis for remote X-ray tuning of graphene transport properties and X-ray sensors comprising the graphene/oxide interface as an active layer. One of the most appealing graphene attributes is the possibility of controlling the type and concentration of charge carriers via application of an electrostatic potential between a grounded graphene layer and a gate electrode, so called gate voltage. The high intrinsic charge carrier mobility in graphene implies a high application promise for use of gated graphene devices – graphene field effect transistors (GFETs) – as high speed electronic devices7. Although a direct application of GFETs in electronic circuits is largely hampered by the lack of a bandgap in a single-layer graphene8, there are advanced devices that do not require the bandgap for their functionality. In particular, graphene spintronic devices9, gas sensors with sensitivity down to the single molecule limit11, and photodetectors12 show great potential for future applications.Graphene has attracted an enormous attention for its unique mechanical, optical, and electronic properties with a wide technological perspective13, graphene-adsorbate charge transfer, molecular orientation, and magnetic properties naming only the most prominent14. However, the possible effect of ionizing X-ray radiation on the GFET properties should be considered. In this paper we show that the X-ray radiation induces strong changes in graphene transport properties via charging of intrinsic defects in the gate dielectric.The research and development of these devices is intimately connected with analysis of their structural, chemical and optical properties. In this respect, the characterization tools based on X-ray radiation are invaluable to determine bond specific chemical composition15. More particularly, defects within the dielectric layer behave like charge traps, which can be ionized, e.g., by electron or hole injection or X-ray radiation, rendering FET sensitive to ionizing radiation16. The effect of charged impurities and adsorbates is even more pronounced in graphene devices18. Recently, the photo-induced doping of graphene has been realized by visible or UV radiation exposure of GFETs26. Here, two distinct groups of GFET devices were introduced: in the first group the charges excited within the photoabsorbing medium  are transferred to graphene appearing as an increase of the graphene DC conductivity22. Within the second group the UV/Vis radiation ionize donor-like traps leaving the gate dielectric positively charged. This charge acts as a positive gate: it increases the electron concentration in graphene by capacitive coupling26. In contrast to direct graphene doping from adsorbed species causing also a decrease of carrier mobility27, the major advantage of “remote gating” is its minimal impact on the charge carrier mobility28.As the semiconductor field effect transistors (FETs) comprise the heart of a modern electronic industry the huge amount of work has been devoted to understanding their properties with respect to their further development. The quality of the gate dielectric has a profound impact on the long term stability of FETset al. studied the effect of the gate voltage on the shift of the core-level peaks associated with graphene and gate dielectric using X-ray photoelectron spectroscopy29. In a separate work, in pursuit for graphene application as an X-ray sensor, Cazalas et al. observed the change in graphene source-drain current of GFET upon hard X-ray irradiation (15 keV) of graphene on SiC30. While the latter work introduces the GFET as a device sensitive to X-ray irradiation, the basic description and understanding of the X-ray radiation effect on the GFET is still missing. Here we show that X-ray radiation induces the ionization of donor-like defects in the gate dielectric leading to a large increase of the electron concentration in graphene, i.e., to its strong negative electrostatic doping (n-doping). This results in an observed shift of the position of the charge neutrality point (CNP) to the large negative values. The succeeding irradiation and simultaneous application of negative gate voltage induces backward shift of the CNP as a result of the compensation of positively charged defects by photoexcited electron current in the gate dielectric, an effect which was not demonstrated for visible or UV irradiation so far.Surprisingly, only little attention was paid to reveal the influence of X-ray radiation on graphene in the GFET configuration. In this respect, Copuroglu 2 gate dielectric and Si substrate beneath as schematically shown in Fig. 31. Subsequently, the graphene was transferred on a p-doped silicon substrate with a 285 nm thick SiO2 layer and pre-fabricated Au/Ti electrodes. Two kinds of devices were used in this study – open and passivated ones. The passivated device comprised a 25 nm thick Al2O3 passivation layer covering the whole device surface whereas the open are left without the passivation layer. The completed GFET was inserted in an ultrahigh vacuum (UHV) chamber. The exposure to X-rays was carried out by Al Kα radiation generated by a standard X-ray source used for X-ray photoelectron spectroscopy. Both types of the devices showed the qualitatively same type of behavior unless explicitly specified.The effect of X-ray radiation was studied employing a graphene field effect transistor with a conventional structure consisting of SiOBG = 17 V. Hence, at the zero external bias the graphene layer is positively doped with holes being the majority charge carriers. For VBG larger than 17 V, electrons become the majority charge carriers. In the following we first describe an initial X-ray irradiation, which is fundamentally different from the subsequent ones and then we turn our attention to these “succeeding irradiations”.The resistivity of passivated graphene as a function of applied back-gate voltage  of the GFET device measured before X-ray irradiation is shown in Fig. BG as depicted in the inset of Fig. BG = −47 V. The subsequent turning of the X-ray source off portrays the CNP shifted to even larger negative values  only after a 24-hour long exposure of the sample to ambient conditions, which is consistent with the UV/Vis irradiation induced doping effect reported to be stable under vacuum at least for several days28.When the GFET is exposed to X-ray radiation at zero back-gate voltage we observe dramatic changes in graphene resistivity in time. As documented in Fig. ues Fig. , thus in26, the permanent change of resistivity upon initial X-ray irradiation can be explained by an accumulation of positive charge within the gate oxide, which via capacitance coupling induces negative graphene doping  interface. Since electrons possess significantly higher mobility in SiO2 than holes they reach either the graphene or the gate on the picosecond time scale whereas the holes diffuse relatively slowly through the oxide layer . Importantly, the SiO2 gate dielectric possesses a significant number of donor-like defects 2 interface33. Donor-like defects are neutral when they are occupied with electron and can be positively charged when one electron is released . These defect sites can be directly photoexcited to create charged defects  we can extrapolate the CNP position to ~−130 V. From this value we can determine the concentration of charge carries in graphene; this is equal to the concentration of charged defects 13 cm−2 taking into account the direct capacitance coupling of trapped charge in oxide with graphene charge carriers.Consistently with the recent reports on UV light induced graphene dopinging Fig. . Here, tBG electrons are driven out of the near G/SiO2 interface region. Hence, in region depleted from electrons there is a lower recombination rate of diffusing holes and, consequently, a higher number of holes approaches this region. The hole diffusion to this region is further promoted by direction of applied external electric field33. Both these effects will result in higher hole capture rate and a faster build-up of the positive charge within the oxide and its higher saturated concentration BG < 0, the electric field drives the electrons towards the G/SiO2 interface while the holes move in an opposite direction resulting in the exactly opposite effect. These effects are documented by observed increase (decrease) of the n-doping rate and saturated graphene doping level if a positive (negative) gate bias is applied , the system can be driven out of the equilibrium between BG = −70 V, the GFET resistivity rapidly increases  a partial decrease of the potential barrier and (ii) its transformation into a potential well  and the CNP shifts back only to −48 V instead to 5 V. If we consider that the charge in the SiO2 is fully compensated, then the CNP position at −48 V is determined by positive charge localized within the non-gated Al2O3 passivation layer. The associated defect concentration 12 cm−2; 2 after the initial irradiation therefore amounts to 6 × 1012 cm−2.The distinct behavior of open and passivated devices originates from the absence/existence of non-gated dielectric near graphene layer, i.e., Al33. To reveal the character of the involved defects we have measured the GFET response to the X-ray radiation as a function of time. The time traces measured on passivated devices for negative VBG are shown in Fig. 2  this gives a significant reduction of the 2 to 2 × 1012 cm−2. The associated potential barrier for the electron flux to graphene drops to ~0.5 eV, which is comparable to the value enabling electrons to flow to graphene. The positive VBG and simultaneous irradiation causes an additional significant shift of the CNP to larger negative values for both open and passivated devices . However, one should take this value with care as it was determined from extrapolated data.In addition to the charge compensation, the electrons can be directly captured by the positively charged donor-like defects  when the VInterestingly, when the succeeding X-ray irradiation was interrupted and after a short period of relaxation renewed, the resistivity quickly increases to the value measured before the turning the X-ray source off and continues as if there was no interruption Fig. . This is26. Since the remote doping does not impose degradation of the charge carrier mobility28, this approach possesses great potential in device tuning. We report on the induced doping level on standard SiO2 substrates that is much higher than has been observed for the UV doping of G/SiO2 or G/h-BN substrates28 and comparable both to chemical doping and UV-induced doping employing highly defective TiOx dielectric26.In this paper we have demonstrated that the X-ray irradiation of graphene field effect transistors causes a strong negative doping of the graphene layer as a result of charging of donor-like defects in the gate dielectrics similarly to recently reported graphene doping by visible and UV radiation2 interface volume, thus neutralizing the built-in positive charge rendering the graphene doping level in GFET devices close to intrinsic one.The strong sensitivity to X-ray irradiation has serious implications for long term stability of GFET devices and GFET characterization employing X-ray radiation as a probe. Concerning the later, the advanced strategies should be utilized to obtain the correct results from X-ray based measurements. In this context, a slight negative voltage and continuous sample irradiation will induce the electron accumulation in the near G/SiO31. Before the growth, the copper foil was heated up to 1000 °C in a home-built reactor and annealed for 30 minutes in a flow of hydrogen . The graphene layer was subsequently grown under the methane atmosphere  for 30 minutes. After cooling the sample to the room temperature, the graphene layer was removed from the bottom side of the copper foil by oxygen/argon plasma treatment .Polycrystalline graphene layers were grown by a standard low-pressure CVD method on ultrasmooth copper foil prepared by the template stripping method35, the graphene was transferred to a p-doped silicon substrate (resistivity of 0.00010–0.0015 Ω cm) covered with a 285 nm SiO2 thin film  and pre-arranged Au (80 nm)/Ti (3 nm) electrodes fabricated by e-beam lithography. Sputter deposition of metal electrodes before graphene transfer is an important step for fabrication of functional GFET structure. The metal contacts underneath graphene possess rough edges from lift-off process ensuring good graphene adhesion and electrical contact. The prepared structure was inserted into a chip holder and contacted by an ultrasonic wire bonder. The breakdown voltage of the dielectric layer (in full device configuration) in our experiments was higher than 4 MV/cm and leakage current through the oxide at VBG = 80 V was lower than 2 nA. The number of adsorbates attached to the graphene surface was decreased by vacuum annealing of the sample at 150 °C for 1 hour in an atomic layer deposition system , which was subsequently used for deposition of a 25 nm-thick Al2O3 passivation layer at the same temperature. The Al2O3 layer was deposited from Trimethylaluminum 3) and H2O precursors employing Ar as a carrier gas  chamber equipped with a manipulator providing five contacts to the sample: one used for the gate, two for the source and drain electrodes and the last two for sample heating.Using a PMMA-assisted wet transfer method−7 Pa) in two terminal geometry, which is sufficient for precise determination of charge neutrality point position. The resistance R of the graphene channel between the metal electrodes was measured using a lock-in amplifier SR830 (Stanford Research Systems) with a frequency of 1333 Hz and a fixed current of 100 nA. The graphene resistivity ρ was calculated from the channel geometry by the formula ρ  = RW/L where W is the channel width (400 μm) and L is its length (50 μm). According to the parallel-plate capacitor model, applying the back gate voltage VBG to a silicon substrate results in a change of the charge carrier concentration in graphene by n = (ε0εr/ed)VBG where ε0 is the vacuum permittivity, εr is the relative permittivity of SiO2 (3.9), e is elementary charge and d is the thickness of the SiO2 thin film. Measuring the graphene resistivity as a function of the back gate voltage (BG trace) then gives us an information about graphene doping and charge carrier mobility. Back gate voltage applied to the silicon substrate was changed during the measurements of BG traces with a sweeping rate of 0.7 V s−1.Electrical transport measurements were performed under the UHV conditions  attached to custom built UHV chamber standardly used for X-ray photoelectron spectroscopy was employed. The photon flux over the sample area was 10Supplementary Information


We report on an optimization of the commonly used wet transfer technique for large-area graphene, grown via chemical vapor deposition, onto aluminum nitride (AlN), which is mainly used as an active, piezoelectric material for acoustic devices. Today, graphene wet transfer is well-engineered for silicon dioxide (SiO2). Investigations on AlN substrates reveal highly different surface properties compared to SiO2 regarding wettability, which strongly influences the quality of transferred graphene monolayers. Both physical and chemical effects of a plasma treatment of AlN surfaces change wettability and avoid large-scale cracks in the transferred graphene sheet during desiccation. Spatially-resolved Raman spectroscopy reveals a strong strain and doping dependence on AlN plasma pretreatments correlating with the electrical conductivity of graphene. In our work, we achieved transferred crack-free large-area (40 × 40 mm2) graphene monolayers with sheet resistances down to 350 Ω/sq. These achievements make graphene more powerful as an eco-friendly and cheaper replacement for conventional electrode materials used in radio frequency resonator devices.The two-dimensional and virtually massless character of graphene attracts great interest for radio frequency devices, such as surface and bulk acoustic wave resonators. Due to its good electric conductivity, graphene might be an alternative as a virtually massless electrode by improving resonator performance regarding mass-loading effects Its higbstrates ,10. Sheeoducibly . Transfeuppliers , which puppliers ,14, but In this publication, we show an optimized wet transfer technique on AlN. As a two-dimensional conductive material, graphene can play an important role as an alternative electrode material regarding the elimination of viscous losses due to its virtually massless character. Using the wet transfer technique , the des2) as a target substrate is highly hydrophilic, therefore, no other surface treatment than standard substrate cleaning is necessary for a successful transfer [SR ≈ 350 Ω/sq as a best value, which is strongly comparable to commercially available graphene on SiO2.Commonly used silicon dioxide (SiOtransfer . Howevertransfer  using grtransfer . Additiotransfer . Our inv2 does not show cracks after transfer under the same transfer conditions (a). In order to investigate surface effects, SiO2 on Si and AlN on Si were characterized via atomic force microscopy (AFM). 2 surface (a) is explicitly smoother than the AlN surface (b). The calculated root mean square (RMS) as a surface roughness parameter averages between 0.9 ± 0.43 nm and 3.49 ± 0.88 nm for SiO2 and AlN, respectively , AlN after an Ar plasma. 2 plasma pretreatment. The effect of N2 and O2 (not shown) is fairly similar as the one for Ar. Comparing the corresponding roughness parameters we obtained RMS values for N2, O2 and Ar (1.1–1.2 nm), which are close to the one for SiO2 (0.9 nm)  measurements were performed. According to the sessile drop method  very sma(0.9 nm) i. For th2, O2 and Ar plasma pretreatments. In general, the contact angle between a fluid and a solid is described by the Young equation [r, which is the ratio between the real and the projected solid surface area [2, O2 and Ar physically damage the AlN structure and smoothen its surface, whereas H2 has almost no major physical impact on AlN due to its low atomic weight. The surface damage is clearly seen in Corresponding CA measurements show a strong change in surface properties for the differently treated AlN samples. The obtained CA a for meaequation :(1)γSA−γace area . The WenApart from this physical smoothing which influences the substrates wettability, there is another important effect responsible for the surface properties. Plasma treatment also leads to a change in the chemical composition of the surface. For all plasmas there is a cleaning effect of the surface. Contamination atoms like carbon and other carbonic composites, which tend to be hydrophobic, are both physically and chemically removed. These composites can readsorb on the AlN surface after the plasma pretreatment , which i2 plasma, there is an oxidation effect of the surface which in turn becomes more hydrophilic due to a smoothening. A certain amount of residual O2 within the reactor chamber might enable oxidizing reactions which results also in a smoothening of the AlN surface. A nanolayer of aluminum oxide allotropes directly increases the wettability. Depending on the fabricated application one needs to keep in mind that oxidation might cause undesired side effects on the device performance 3 and NH4NO3, due to the aforementioned, non-negligible amount of residual oxygen in the plasma chamber. Despite their negligible effect in physical surface smoothening, H2 atoms might create labile hydroxide compounds with the assistance of background oxygen, such as Al(OH)3. These chemical reactions can also activate the AlN surface due to the availability of aluminum atoms in the substrate material. The creation of nitrate components as intermediate products of chemical reactions increase, as well as oxides, the surface energy i.e., wettability [Physical damage in combination with Ntability . Nitrateθ* for all applied plasmas within the first minutes after plasma exposure. This drop in CA is rapidly recovered within one hour after pretreatment. The recovery effect is quickest for H2 plasma. There is a specific saturation CA value for each of the plasma pretreatments, which is time-independent and averages between 44° for H2 and 18° for N2, O2 and Ar (dashed lines). Hence these maximum values of t > 2 h for different plasmas represent the ones shown in 2, and O2 plasma several oxidation reactions superpose the cleaning effects, whereas for H2 plasma a stable oxidation apparently does not take place. For this purpose a profound analysis of the chemical compounds on the AlN surface after plasma treatment needs to be carried out, this is not within the scope of this publication.The “cleaning” phenomenon of the removal of surface contaminations can be seen in θ* < 20° improves the quality of transferred graphene. The Raman D/G ratio decreases below 0.2 and the crack density can be lowered significantly. As shown later on this influences directly the graphene sheet resistance.In a first summary we can conclude that plasma pretreatments impact the surface both physically and chemically, whereas a chemically activated surface shows an aging effect. Transfer experiments indicate that already a physical impact on the AlN surface resulting in as little as 2 and AlN on Si are shown as a reference. Although the two samples differ in their surface roughness (RMS = 4.4 ± 0.7 nm (1.8 ± 0.2 nm) for B1 (B2)) the CA is remarkably high for B2 (63 ± 6°). When an Ar plasma is applied on both samples RMS values do not change  line shows different levels of hole doping (strain). The slopes are obtained from [Additionally, the strain distribution across the transferred graphene layer was investigated to further understand the effect of plasma pretreatment. Using spatial Raman spectroscopy analysis of the 2D and G peak positions, ω2D modes . In Figuned from , as(3) would not hold. Hall measurements of graphene on AlN showed hole doping, which allows the use of the aforementioned equations. Das et al. ,25 revean et al.  calculatD2 versus ωG reveals a significant change for plasma pretreated AlN on Si. Strain and doping levels are labelled in 2 and Ar plasmas, whereas there is no dependency identifiable for untreated AlN and AlN pretreated with an H2 plasma. Graphene on Ar (O2) plasma pretreated AlN shows dominantly tensile strain between 0.1–0.3% (0.1–0.2%); the doping is significantly lower for the Ar plasma (n = 2 × 1011–1 × 1013 cm−1) compared to the O2 plasma (n = 2 × 1013–4 × 1013 cm−1). The same result is obtained for N2 (not shown). We can conclude that variations of strain and doping are strongly reduced applying an N2, O2 or Ar plasma instead of an H2 or no plasma pretreatment at all. To obtain a greater spatial distribution map measurements were done, shown in (e–g) for untreated and Ar plasma treated AlN substrate (AlN/Si and B1). Due to observable D peak intensities for graphene on untreated AlN (e) information about actual doping concentrations cannot be extracted due to overlapping effects in the Raman shifts. However, a clear reduction of strain variation is apparent for the Ar plasma for both AlN/Si and B1 . The same results hold for B2 (not shown). This indicates that the transfer-induced strain results not from surface roughness itself but from wettability effects on the graphene sheet during transfer. For the transfer of graphene on B1 we added exemplary AuCl3 during the Cu etching, which results in an additional doping of the transferred layer. This is directly visible in a shift of the 2D/G ratio (g). AuCl3 might play an important role regarding the decrease of sheet resistance due to the doping effect. This will be investigated further.In −1) vanishes almost completely, whereas the 2D/G intensity ratio increases up to 2.6. The 2D peak lies at 2772 cm−1. Both indicate the existence of mostly monolayer graphene on AlN [In order to quantify the enhanced graphene quality due to the applied plasma treatments the samples were investigated via optical microscopy and Raman spectroscopy. SR, shown in SR constantly reaches values below 1 kΩ/sq. Best values were obtained as SR = 350 Ω/sq. Sheet resistance measurements were homogenous on an area of 40 × 40 mm2. So far, we can exclude the effect of induced additional strain due to doping, as shown in SR seems to be promising. Additional experiments on this topic still need to be carried out.The obtained quality improvement is directly proven via four-point measurements. Due to plasma pretreatment, the sheet resistance 2 (1)) ensure a continuous desiccation via capillary forces in between the substrate and the graphene/PMMA (Poly (methyl methacrylate)) whereas hydrophobic surfaces (such as AlN (2)) form water droplets instead. These droplets either remain in between the substrate and the graphene layer (1) or they break the graphene layer for sublimation, creating cracks and discontinuities (2). As shown experimentally in this work, additional strain is applied on the transferred graphene sheets, which critically influences the sheet resistance SR of the graphene. The underlying effect of hydrophobic surfaces is the concept of surface and interfacial energies, which we identified indirectly via CA measurements. Hydrophobic surfaces feature a low surface energy and hydrophilic surfaces feature a high surface energy [SR. Problems regarding the transfer of graphene onto graphene will occur due to its extremely hydrophobic behavior. CA measurements revealed θ* to be very close to 90° .In this work, the synthesis of a graphene monolayer was achieved via chemical vapor deposition (CVD), a mature and commonly applied technique of growing graphene , on copp2 pieces for 10 × 15 mm2 graphene transfers. Solidly mounted BAW resonators (BAW-SMR) with sputtered AlN as a piezoelectric layer  and 1.5 µm AlN on MoAlN (B2)) were used for 40 × 40 mm2 graphene transfers ). The AlN wafers were cut into 20 × 20 mmransfers . Both Al−1 clearly identifies this as a monolayer structure [The graphene on Cu foil was routinely investigated by a Raman spectroscopy line scan of 121 data points , since it is an approved, easily applicable, non-destructive characterization method . The reptructure ,31. An a4)2S2O8, 0.5 mol/L). This turned out to result in a cleaner interface between AlN and graphene after transfer due to lower formation of precipitates and chemical byproducts than commonly used etching solvents like iron chloride (FeCl3) and iron nitrate (Fe(NO3)3). Wu et al. [4)2S2O8 as an etching solvent since it avoids additionally induced Fe compounds during etching, which reduce the quality of the grown graphene. The PMMA/graphene stack was then deposited onto different AlN target substrates. Before the graphene transfer, the AlN substrates were exposed to different plasmas  in order to vary the surface conditions regarding surface energy and roughness. The SiO2 substrates received no plasma treatment and were used as reference samples. For comparison purposes, graphene transferred on untreated AlN was also investigated. Plasma pretreatments of AlN samples were done in a direct current (DC) plasma reactor . The wettability of the AlN sample was characterized via contact angle measurements using the sessile drop method [After CVD synthesis, the graphene monolayers were transferred on differently pretreated AlN substrates using the wet transfer technique , describu et al.  suggest p method  in a sel2, O2 and Ar plasma pretreatments. Focusing on the chemical “cleaning” of the surface leads to a temporal but strong increase of wettability, which enables a defect-free graphene transfer without changing the AlN’s surface morphology. Sheet resistance measurements show a direct effect of this optimization regarding sheet conductivity. SR can reproducibly be reduced below 1 kΩ/sq for transferred graphene with the size of 40 × 40 mm2 and an Ar plasma pretreatment. The wet transfer of CVD graphene monolayers onto AlN was investigated. We identified a strong dependence of the wettability of AlN surfaces on different plasma pretreatments due to both a physical and a chemical effect on the AlN surface. AFM images and contact angle measurements reveal a change of surface properties resulting in spatially lower strain variation and a tremendous reduction of layer cracks and gaps for NWith this new knowledge, the wet transfer method for the assembly of multilayer graphene sheets for further sheet resistance reduction should be investigated further. A sensitive adjustment of the surface properties of graphene regarding its strong hydrophobic behaviour is essential to obtain even lower sheet resistances. Furthermore, the effect of substrate surface pretreatment is an interesting and inevitable topic for the transfer of high quality graphene onto other substrate materials. Our work clearly shows a strong improvement in the quality of graphene and therefore emphasizes the promising future of graphene as a virtually massless and ultrathin electrode material for high-frequency devices.