However, the most important advantage of the introduced metallic alloy-decorated graphene is its distinct stability in acid medium; the retention in the electrocatalytic performance after 1,000 successive cycles is approximately 98%. This finding is attributed to the high corrosion resistance of the NiFe alloy. The kinetic study indicates that the number of the transferred electrons is 3.46 and 3.89 for the introduced and Pt/C (20 wt%) electrodes, respectively which concludes a high activity for the proposed nanocomposite. The suggested decorated graphene can be synthesized using a multi-thermal method. Typically, nickel acetate, iron acetate, graphene oxide and urea are subjected to MW heating. Then, sintering with melamine in an Argon atmosphere at 750 °C is required to produce the final electrocatalyst. Overall, the introduced NiFe@ N-doped Gr nanocomposite shows remarkable electrochemical activity in the acid medium with long-term stability.NiFe nanoparticles-decorated & N-doped graphene is introduced as an effective and stable non-precious electrocatalyst for ORR in the acid medium. Compared to conventional Pt/C electrodes under the same conditions, the proposed nanocatalyst shows closer onset potential and current density. Typically, the observed onset potentials and current densities for the synthesized and Pt/C electrodes are 825 and 910 mV (vs. NHE) and −3.65 and −4.31 mA.cm Undoubtedly, the confirmed near depletion of fossil oils is forcing the researchers to find effective, low cost, and environmentally safe energy devices. In addition to this dilemma, the serious negative impact to the climate promotes the elimination of the fossil fuels utilization. Flexibility, ease of management, and applicability make fuel cells the most promising energy devices to overcome the aforementioned problems. For example, wide utilization of the proton exchange fuel cells (PEMFCs) was one of the closing remarks of the 2015 Paris Climate Conference (COP21).1. Although there are numerous efforts to develop new membranes for the alkaline media fuel cells2, commercially, the acidic solutions-based PEMFCs are prevailing due to the excellent performance of the Nafion membrane. However, working in an acid medium limits the selection of the electrode materials as most of the metals show low corrosion resistance to acids. Accordingly, precious metals  became inevitable choices for manufacturing the commercial electrode materials3. In acid media, platinum (Pt) is the most effective catalyst for both hydrogen oxidation and oxygen reduction5. In addition to Pt, palladium-based compounds also have considerable activity7. Though, the high cost and rarity of the precious metal electrodes are constraining the wide application of the PEMFCs8.The main constituents of any PEMFC are anode, cathode, and membrane. Typically, oxygen is reduced at the surface of an active cathode while the fuel  is oxidized at the anode surface9. Recent research has indicated that bimetallic nanoparticles containing a first-row transition metal possess magnetic, optical, composition-dependent and catalytic properties that are different from mono metallic nanoparticle components10.Disregarding the stability, many non-precious materials have shown good performance as anodes11 and CNT/graphene mixture12), supporting of effective bimetallic nanoparticles on a proper N-doped carbonaceous material can distinctly enhance the ORR electrocatalytic activity13. Besides enhancing the electrocatalytic activity, N-doped carbon nanostructural supports showed more stability than nitrogen-free ones due to the high number of surface nucleation sites, which allows for anchorage and high dispersion of the catalyst nanoparticles on surface of the support material16. Moreover, due to the strong electron donor behavior of nitrogen, the doping process improves the durability of the produced carbon-support catalysts because of enhancement of π bonding18 and their basic properties19.Considering that the most effective non-precious ORR catalysts are primarily nitrogen-doped nanocarbons (e.g. N-doped carbon nanotubes (CNT)2 g−1), strong mechanical strength, and excellent electrical conductivity. Consequently, it was exploited to enhance the performance of several promising electrode materials in the electrochemical devices22. Moreover, compared to other carbon nanostructures, the chemical route for graphene synthesis provides a good chance for functionalization by active groups which aids in decoration surface by metallic nanoparticles9.Graphene is a charming carbon material with excellent characteristics including a large theoretical surface area  Exploiting the alloy structure of the transition metals to enhance the stability in the acid media. (2) Maximizing the nitrogen content in the carbonaceous support to strength the catalyst activity toward the ORR. (3) Synthesis of the graphene sheets supports by the chemical route to develop chemical anchors for the transition metals nanoparticles which strongly improves the attachment and consequently ameliorates the activity of the final catalyst. Overall, the obtained results indicated that the proposed nanocomposite have very good electroactivity toward ORR with distinct stability in the acid media.25. However, after the violent oxidation, the graphite peak disappears and a new diffraction peak appears at 2θ of 10.5° 25. On the other hand, due to the reduction process, the reduced graphene oxide shows a broad peak that can be fitted using a Lorentzian function into three peaks, which are centered at 2θ = 20.17°, 23,78° and 25.88°, which correspond to the interlayer distances of 4.47, 3.82 and 3.53 Å, respectively. These XRD results are related to the exfoliation and reduction processes of GO and the processes of removing intercalated water molecules and the oxide groups27. The observed broaden peak indicates the smaller crystalline size of graphene in the single-layer or few-layer structure. Accordingly, from left inset , iron and nickel are identified by the standard peaks at 2θ of 44.67°, 65.02° and 82.33°, and 44.05°, 54.85° and 76.37° corresponding to (110), (200) and (211), and (111), (200) and (220), respectively which indicates that some nanoparticles have a physical Fe/Ni mixture. Additionally, no peaks attributed to oxides or carbides were detected. According to the utilized characterizations which confirmed formation of pure Fe and Ni and considering the difficulty of evaporation of these metals due to the high melting points , the weight of metals in the final produce can be estimated. Moreover, it was reported that calcination of the graphene oxide prepared by a similar chemical route to 750 °C leads to lose around 60 wt%29. Consequently, the Ni:Fe:C ratio in the produced composite can be determined to be 41:40:19 wt%, respectively.Accordingly, the prepared GO was used to prepare the proposed FeNi-decorated & N-doped graphene as it was explained in the experimental section. The XRD pattern of the synthesized composite is displayed in the left inset Fig. . The obs30.Although XRD is highly trustable analytical technique, its utilization is limited to the crystalline materials. Therefore, to investigate nitrogen doping, X-ray photoelectron spectroscopy (XPS) was exploited. The obtained XPS spectra Fig.  indicateFigure −1 scan rate and room temperature. The synthesized electrode reveals comparable performance with the precious metal electrode in the form of current density and onset potential. Based on previous reports, the onset potential is defined as the potential at which the background subtracted current density is equal to 0.1 mA.cm−2 32. The onset potential for the introduced decorated graphene was close to that of Pt/C ), Fig. 19, while the metallic nanoparticles have more adsorption capacity for the molecular oxygen. Delivery of electrons, from the anode to the cathode, leads to complete the oxygen-hydrogen combination to form water molecules; ORR reaction. From the chemistry point of view, the pyridinic nitrogen has more basic characteristic than pyrrolic because the nitrogen lone pair electrons do not share in the pyridine cycle resonance. However, in case in pyrrole, the lone pair electrons contribute in the cycle resonance which negatively affects the basic characteristic. Accordingly, it is expected that pyridinic nitrogen has more attraction capacity for the hydrogen ions that reflects more contribution in the ORR reaction. It is noteworthy mentioning that the expected adsorption influence of the utilized N-doped graphene support can partially enhance the electrocatalytic activity of the proposed composite. However, the main impact can be assigned to avoiding the agglomeration of unsupported bimetallic nanoparticles which strongly improves a very important parameter in the heterogeneous catalytic reactions; contact area between the reactants and the catalyst surface. Furthermore, the excellent electrical conductivity of the utilized support has also a regarded effect. The proposed mechanism of O2 reduction on the surface of the introduced catalyst is visualized in Fig. Doping the carbon nanostructures with heteroatoms such as N can distinctly change the properties of carbon. For instance, doping of carbon by nitrogen strongly enhances the oxidation resistance capability and its ORR catalytic activity. For example, doping of carbon nanofibers by nitrogen led to increase the onset potential of the oxygen reduction reaction by 70 mV with a corresponding electron transfer number of approximately 437. The stability of the introduced and the precious (Pt/C) electrodes was first investigated by cyclic voltammetry analysis for 1,000 successive cycles. Figure th and 850th cycles of the introduced FeNi-N-Gr  of 1.23 V is difficult to be obtained. Instead, the steady-state rest potential of the platinum electrode in the oxygen-saturated solutions is around 1.06 V, a mixed value of the thermodynamic potentials of Pt/PtO and On) in the ORR is another important factor. Rotating ring-disk electrode (RRDE) analysis is typically invoked to measure the number of the utilized electrons. This technique can give indication about the relative importance of the H2O2 routes in the overall oxygen reduction reaction process. However, in the literature, for the same material, several n values can be found. These different values of the number of transferred electrons can be attributed to the history of the electrocatalytic material which can distinctly affect the ORR rates. For instance, at the ring, in addition to the oxidation of hydrogen peroxides that generates an anodic current, decomposition of H2O2 without current flow may take place. Moreover, the Pt ring potential can influence the number of electrons, e.g., by varying the relative amount of the formed PtO, and also the synthesis conditions of the investigated material can play a strong role. Therefore, other techniques have been introduced to estimate the number of electrons, such as scanning electrochemical microscopy41 and cyclic voltammetry42. In the cyclic voltammetry-based procedure, the analysis is performed at different scan rates and the peak currents (Ip) increase linearly with the scan rate that is a typical characteristic of the reaction occurs on the surface of the electrode. The number of the electron can be estimated from the slope according to this equation:n is the number of the electron transfer, A is the electrode active area (0.073 × 10−4 m2), v is the scan rate (mV/s), and S is the concentration of the adsorbed oxygen on the electrode surface . From the Ip vs. v linear relation slope, the number of electrons n can be calculated. The voltammograms for the FeNi-N-Gr and Pt/C electrodes are introduced in the Fig. R2 = 0.998) for the FeNi-N-Gr electrode data. Accordingly, the number of electrons was determined to be 3.89 and 3.465 for Pt/C (linear regression is not shown) and the introduced electrode, respectively.In addition to the current density and onset potential, the number of sharing electrons , −4 M) at 20 °C, −9 m2.s−1), υ is the kinematic viscosity of the solution at 25 °C (1 × 10−6 m2.s−1). As can be distinguished from the value of the linear regression parameter (R2 = 0.994), the experimental results strongly validate the model, moreover the high linearity obtained suggests first-order of the reaction kinetics toward the concentration of dissolved oxygen43. The estimated number of electrons from eq. ; Fig. In summary, graphene sheets were decorated with FeNi nanoparticles by hydrothermal treatment of graphene oxide in the presence of nickel acetate and iron acetate. Moreover, the presence of urea in the reaction medium followed by calcination in an argon atmosphere leads to the incorporation of nitrogen atoms in the graphene skeleton. The produced FeNi-decorated and N-doped graphene can be exploited as a stable and effective electrocatalyst for the oxygen reduction reaction process in acid medium. The high performance was attributed to the good affinity of the metallic nanoparticles and nitrogen atoms to oxygen molecules and hydrogen ions, respectively. Moreover, the good stability in acidic media can be assigned to the alloy structure of the metallic nanoparticles.2SO4 (assay 95–97%); these chemicals were purchased from Sigma–Aldrich. All the used chemicals were utilized without further modification. DI water was used as a solvent.In the introduced decorated graphene, the used precursors for the metal nanoparticles were iron (II) acetate , and nickel acetate tetrahydrate . The reduced graphene oxide was prepared by chemical route using graphite powder (particle size <20 μm), hydrazine monohydrate, hydrogen peroxide, and H44. Briefly: treated twice by 5% HCl five grams of graphite was placed in ice bath concentrated H2SO4 (130 mL). Later on, 15 g of KMnO4 was added gradually to the mixture under zero Celsius temperature condition with stirring for 2 h. Then, distilled water was added to the mixture which results in increasing the temperature to 98 °C. After that the mixture was cold to room temperature, and H2O2  was added, the mixture was kept under stirring for 24 h. Later on, the synthesized GO was separated by filtration under vacuum and washed by10% aqueous HCl several time and then dried at 50 °C. NiFe alloy nanoparticle-decorated and N-doped graphene was synthesized from mixing 0.5 mM iron (II) acetate and 0.5 mM nickel (II) acetate tetrahydrate aqueous solutions with 250 mg of urea (as a source of nitrogen) and stirring for 2 h, followed by ultrasonication for 30 min. As maximizing the nitrogen content in the final product was an important target during the synthesis process, using of urea during preparation of the FeNi-decorated graphene was done (in the reflux step) to incorporate nitrogen atoms within the graphene cycles which effectively increases the nitrogen content during the thermal treatment with melamine45. It is worth mentioning that, based on our studies and others, acetate salt was chosen due to the complete reduction during calcination at relatively high temperature under inert atmosphere to produce the corresponding metals rather than the expected metal oxides47. In another beaker, two hundred mg of the prepared graphene oxide were treated in a microwave oven for two minutes at around 600 W to achieve the thermal exfoliation. The two solutions were mixed and the obtained slurry was then refluxed for twelve hours at 150 °C. The produced slurry was filtered, and the solid material was dried for one day at 80 °C under vacuum. The dried powder was ground with twice as much melamine and calcined under argon atmosphere at 1 atm for four hours at 750 °C. High calcination temperature was chosen to ensure complete reduction of graphene oxide to graphene and avoid formation of metal oxides46. Both urea and melamine were used as nitrogen precursors to enhance the nitrogen content in the final product.The graphene was prepared chemically from reduction of exfoliated graphene oxide (GO). Typically, GO was synthesized from natural graphite powder by a modified Hummer’s method2) of the glassy carbon electrode which was then subjected to drying process at 80 °C for 20 min. Cyclic voltammetry measurements were carried out in 0.5 M H2SO4 solution and the sweep potential range was adjusted from −0.2 to 1.0 V [vs. Ag/AgCl].Information about the phase and crystallinity was obtained by using Rigaku X-ray diffractometer  with Cu Kα (λ = 1.5406 Å) radiation over Bragg angle ranging from 10 to 80°. Normal and high resolution images were obtained with transmission electron microscope  operated at 200 kV equipped with EDX analysis. The electrochemical measurements were performed on a VersaSTAT 4 (USA) electrochemical analyzer and a conventional three-electrode electrochemical cell. A Pt wire and an Ag/AgCl electrode were used as the auxiliary and reference electrodes, respectively. All potentials were quoted regarding to the Ag/AgCl electrode. Glassy carbon electrode was used as working electrode. Preparation of the working electrode was carried out by mixing 2 mg of the functional material, 20 µL Nafion solution (5 wt%) and 400 µL isopropanol. The slurry was sonicated for 30 min at room temperature. 15 µL from the prepared slurry was poured on the active area (0.073 cm


We present Raman studies of graphene films grown on copper foil by atmospheric pressure CVD with n-decane as a precursor, a mixture of nitrogen and hydrogen as the carrier gas, under different hydrogen flow rates. A novel approach for the processing of the Raman spectroscopy data was employed. It was found that in particular cases, the various parameters of the Raman spectra can be assigned to fractions of the films with different thicknesses. In particular, such quantities as the full width at half maximum of the 2D peak and the position of the 2D graphene band were successfully applied for the elaborated approach. Both the G- and 2D-band positions of single layer fractions were blue-shifted, which could be associated with the nitrogen doping of studied films. The XPS study revealed the characteristics of incorporated nitrogen, which was found to have a binding energy around 402 eV. Moreover, based on the statistical analysis of spectral parameters and the observation of a G-resonance, the twisted nature of the double-layer fraction of graphene grown with a lower hydrogen feeding rate was demonstrated. The impact of the varied hydrogen flow rate on the structural properties of graphene and the nitrogen concentration is also discussed. Due to its high charge carrier mobility , it has e planes –6. For SThe controlled injection of defects, which cause strain, is an extra degree of freedom in addition to the number of layers. This may account for the unique properties of TG. In particular, it was shown that the strained TG bilayer could be an ideal platform for the realization of the high-temperature zero-field quantum valley Hall effect . From a 4) is the most common hydrocarbon used in the CVD process to grow graphene. However, the use of hydrocarbons other than CH4 compounds is a challenging task. The successful implementation may offer the possibility to tune the growth process and to pave the way to graphene synthesis with desirable parameters, such as type of defects and their concentration.TG can be obtained by different methods, e.g., by means of graphene folding, graphene layer stacking, thermal decomposition of SiC  or chemin-decane as a precursor in the presence of nitrogen flow.In this work, we investigate the experimental conditions at which the APCVD growth of large area, nitrogen-doped TG can be realized utilizing 2 sample was placed in the middle of the reactor. One side of the copper foil covered the inner wall of the reactor. The copper foil was annealed for one hour at 1050 °C in the presence of N2 and H2 gas flow at a rate of 100 and 150 cm3/min, respectively. The purity of the nitrogen gas was 99.95%. We utilized a commercial hydrogen generator (GVCh-12D) as a H2 source. The resulting purity of the hydrogen gas was 99.99%. The temperature was controlled by a thermocouple placed inside the heating block, next to the reactor wall.A custom-made APCVD set-up with a 14 mm diameter tubular quartz reactor was employed for the experiment. Polycrystalline copper foil  with a thickness of 60 μm was used as the catalyst. Prior to the APCVD, the foil was electrochemically polished for 5 min in 1 M phosphoric acid at a bias voltage of a 2.3 V. The 35 × 45 mmn-decane. In addition, in previous publications [The above experimental procedure has been previously reported in –14. The ications –14, the 2 and H2 gas flow with the rates of 100 and 60 cm3/min, respectively. Sample B was prepared under similar conditions, except the H2 gas flow rate was reduced to 6 cm3/min.In this article, we present the results related to two samples: sample A and sample B. Sample A was prepared at 1050 °C in the presence of Nn-decane (C10H22), a member of the homologous series of alkane hydrocarbons. n-Decane has a molecular mass approximately one order of magnitude greater than that of methane, which influences the growth kinetics of graphene. As a representative n-alkane, n-decane forms chains of radicals with a high reactive nature during thermal decomposition. This could stimulate numerous chemical reaction pathways and promote doping. In fact, CVD makes it possible to dope graphene by nitrogen in situ, which not only tolerates the ground state of graphene via additional electrons but also introduces a strain to graphene because of the difference in ionic radii [n-decane could lead to the decomposition of the nitrogen molecule, which in fact has one of the strongest binding energies. The resulting atomic nitrogen can be embedded into the graphene lattice.The APCVD depends essentially on the hydrocarbon precursor . To thisic radii . The radn-decane was introduced into the tubular quartz reactor via barbotage system for 30 min. The feeding rate of n-decane was estimated to be 4 μL/min (for both samples). Afterwards, the tubular quartz reactor was cooled at a rate of 50 °C/min in the presence of N2 gas flow. The obtained properties of samples A and B are typical for samples synthesized under similar conditions.The 2 forming gas annealing [2-based annealing [2(598 nm)/Si substrates without the use of a polymer support. This was performed in two steps. First, one side  of the copper foil was treated for 3 min in a solution of H2NO3 and H2O mixed in a volume ratio of 1:3, and then the copper foil was totally dissolved in a water solution of FeCl3. The graphene film was gently washed several times in a bath with distilled water prior to the transfer onto the substrate.The transfer of graphene from the original to the arbitrary substrate without deteriorating the crystallinity of the graphene is still a challenging task . Currentnnealing , O2-basennealing , and in nnealing ), deterinnealing . In this−1 for both spectrometers. A 3D scanning laser confocal Raman microscope (Confotec NR500) allowed for the acquisition of two kinds of images within a single scan: a Rayleigh image, using laser light reflected from a sample, and a spectral image by Raman scattering. More details about the laser beam size and spectra accumulation time are presented further in the text.The graphene samples were analyzed by Raman spectroscopy using the Nanofinder HE with 532 nm and 473 nm excitation wavelengths and a Confotec NR500 confocal micro-Raman spectrometer with 473 nm excitation wavelength. The spectral resolution was about 3 cmhν = 1486.6 eV) was used for X-ray photoelectron spectroscopy (XPS) measurements. The base pressure in the analytical chamber was lower than 2 × 10−7 Pa. The 20 eV and 40 eV pass energy values of a hemispherical electron energy analyzer were used for the survey and high resolution spectra acquisition, respectively. The energy scale of the system was calibrated with respect to Au 4f7/2, Ag 3d5/2 and Cu 2p3/2 peak positions. ESCALAB 250Xi Avantage software was used for the peak deconvolution and fitting procedure using a sum of Lorentzian–Gaussian (70:30) functions. The samples were analyzed as received and no surface cleaning procedure was applied. Finally, the transmittance was measured using the PROSCAN MC-121 spectrometer.A Thermo Scientific ESCALAB 250Xi spectrometer with monochromatic Al Kα radiation  was performed for the same sample area as in I2D/IG  utilizing the same laser beam conditions as for 473 nm excitation. By analogy with the previous Raman maps, the non-monotonic distribution of mapped values was observed. As an example, in the inset of ID/IG ratio is becoming a common method for the point defect concentration evaluation [In many aspects, the theory of Raman scattering in graphene is very well understood and the quantitative analysis of Raman spectra parameters provides sufficient information about structural and electrical properties of graphene. For example, the analysis of the line shape of the 2D band along with its spectral position can provide important information about the number of layers in graphene and the interlayer interaction . From thaluation –29.−1 and 67 cm−1, respectively. This suggests that within the same sample there are at least two graphene ”systems” with quite different numbers of layers. Indeed, the presence of two maxima in the 2D-position distribution  could be caused by a double layer structure of graphene [−1 . The obtained results are in reasonable agreement with each other. Therefore, in order to demonstrate the universality of the elaborated method, for sample B on a SiO2/Si substrate, the Raman data set splitting was based on another criterion, namely, the 2D-band position with a splitting value of 2712 cm−1 . The maxima of the G-band position distribution for the SLG fraction of sample A and B are centered at ≈1590 cm−1  and ≈158−1  [2C peak position is usually explained by the charge transfer phenomena that take place in the substrate–graphene system [Eb can be related to the difference in concentration of sp2C–H bonds which cause the down shift of the Eb[3C bonds [2C–N bonds [In the inset of  Pt(111) –33 to 28 Pt(111) –36. Thise system . Since bof the Eb. The ori3C bonds –40. Fina–N bonds –44. The –N bonds . A numbe–N bonds –38. TherEb = 399.7 eV and 402.1 eV (sample B) (Eb = 401.8 eV (sample A)). The peak at Eb = 399.7 eV observed for both samples is related to the adsorbed nitrogen. Its position was quantified by the measurement on the bare substrate area, see inset of Eb ≈ 402 eV is greater for sample A. We would like to emphasize that these observations are consistent with the Raman spectroscopy results, from which it follows that the G-band blue shift is greater for sample A.Finally, the quantified surface atomic concentrations determined by XPS for samples A and B are summarized in 15 cm−2, the average surface concentration of the nitrogen atoms in sample B can be evaluated as nN ≈ 1.5 × 1013 cm−2.The XPS measurements provide the atomic concentration of nitrogen in the studied samples, as shown in ID/IG provides information about the concentration of point defects n [On the other hand, the Raman integral intensity ratio efects n ,ID/IG ≈ 1, and the energy of the laser Elaser ≈ 2.33 eV, we get n ≈ 2.2 × 1011 cm−2.From the experimental results, nN value may be the following. The nitrogen atoms could be located in such a configuration that almost does not affect the ID value. In particular, the pyrrolic nitrogen due to the symmetry breaking of hexagon rings should have a strong impact on the intensity of the D band and cannot be considered as the main type of defect in the studied films.The reason for lower defect density as compared to the Now we focus on the Raman data associated with the double layer graphene. In For the domains corresponding to the spectra presented in a is the lattice parameter of graphene (2.46 Å),  is the reduced Planck’s constant, vF is the Fermi velocity in monolayer graphene (106 m/s) and Elaser is the laser energy. Within this approach we obtain θ = 13.4° and 11.9° for the laser excitation wavelengths of 473 and 532 nm, respectively. The estimated θ values mean that in the investigated domains, the layers of graphene could be electronically decoupled [where aphene 2.6 Å,  is The higher G band intensities were observed at different locations within the same probed area for different excitation wavelengths. This fact confirms that the presence of the G-resonance is a consequence of the twisted nature of the graphene. However, the ratio of integrated G-band intensities for the double layer TG and SGL is ≈60 and ≈30 for excitation wavelengths of 532 and 473 nm, respectively . In our It is worth highlighting that the above presented Raman mapping has been performed on different sections of samples (not shown here). One of the main results of this study is the uniformity of the samples, confirming their macroscopic homogeneity. For sample B we have provided sets of data where SLG fraction was significant (≈50%), which helps to evaluate and compare the structural properties of SLG and TG. However, for other sets of data, the fraction of double layer TG dominates.We also proved homogeneity directly by the light transmittance measurement of samples transferred on a glass substrate. The diameter of probed area was ≈0.5 cm. In Now we would like to discuss the two most prominent phenomena observed in our experiments: (i) the morphology change of the films with hydrogen flow rate and (ii) nitrogen atom incorporation into graphene, which we believe to be correlated phenomena. Nevertheless, as a first approach, we will discuss them separately.Let us underline the main features that distinguish the morphology of studied samples. Sample A is characterized by the presence of hexagonally shaped domains with a few and single layer graphene domains. In turn, sample B contains the mixture of single and double layer domains where the double layer fraction dominates. Our studies do not indicate the presence of any regularity in shape and distribution of SLG and double layer domains at the measured scale. Since the hydrogen flow rate value was the only technological parameter we have varied during the synthesis, we will focus on the impact of hydrogen on the graphene. First of all, the hydrogen partial pressure strongly affects the shape of the graphene edges. It was demonstrated that higher hydrogen pressure favors the hexagonal shape whereas low hydrogen pressure makes dendritic-like growth preferable –52. Our 10H22 decomposition there is enough hydrogen to terminate the graphene edges [The formation of few layer graphene requires formation of the additional graphene layer (layers) on the top or underneath of a SLG. These processes are usually described by wedding cake (WC) –54 and ine edges .2 gas was used as the nitrogen source during the CVD growth [2 gas as a nitrogen source arises from the fact that nitrogen molecule possesses one of the strongest bonds with an energy of 226 kcal/mol, which means that the temperature of the CVD process is not enough to decompose a significant amount of nitrogen molecules into atoms. The authors of the work [•C) or hydrocarbon (•CxHy) radicals may attack on nitrogen molecules [2 dissociation process. For example, for the •CH + N2 = N + •CHN reaction this value was determined as 21.2 ± 0.7 kcal/mol [n-decane just by thermal decomposition [Finally, we turn to the problem of nitrogen incorporation into graphene. Nitrogen can be incorporated into graphene sheet (i) in situ, using ammonia as a component of the gas carrier mixture  or with D growth . In our the work  did not olecules . The endkcal/mol . Moreoveposition n-decane decomposition at elevated temperature over copper catalyst is beyond the scope of our study. We can only reasonably assume that the different partial pressure of H2 in a gas mixture could favor some decomposition path. As a consequence, it enhances the concentration of radicals with specific configurations. Obviously, the enthalpy of the hydrocarbon reaction with N2 depends on the hydrocarbon radical configuration [However, the issue of the exact reaction route of guration . Therefo•CxHy + N2 reaction has been proposed.In summary, the graphene films have been studied by using the micro-Raman technique. The samples have been grown on copper foil by APCVD using n-decane as a precursor and a mixture of nitrogen and hydrogen as a carrier gas with the use of different hydrogen flow rates. A special analysis approach to the Raman data was employed based on the statistical analysis of spectral line parameters. This approach allowed the association of Raman spectra to fractions of the films with different thicknesses. Based on the values of the 2D peak FWHM and 2D band position, the double layer fraction of graphene grown with the lower hydrogen feeding rate has been established. Moreover, the analysis of the Raman spectra revealed the presence of graphene spots with the G-resonance for both excitation wavelengths used in our experiments (473 nm and 532 nm). The observation of the G-resonance directly confirms the twisted nature of graphene. The obtained blue shift of the G and 2D band positions of the SLG fractions is caused by nitrogen doping, which has been proved by the XPS study. The binding energy of incorporated nitrogen has been evaluated to be around 402 eV. The amount of the G-band shift for each sample is consistent with the XPS data. It has been found that at the wavelength of 550 nm the transmittance for the film grown with the lower hydrogen feeding rate is equal to 94%, which corresponds to 2–3 graphene layers. This is in good agreement with the micro-Raman findings. We suppose that the variation in the morphology is presumably related to the variation of the hydrogen flow in our experiments, as it has been also demonstrated by other authors. Finally, the possible mechanism of the nitrogen concentration incorporated in graphene based on a variation of the endothermicity of a Further work related to the mechanisms of the nitrogen doping and relation between transport properties of graphene and its microstructure is in progress now.


The as-synthesized composite exhibits excellent electrochemical properties by using the advantage of NCS nanontube core as superhighway for electron and ion transport, and CNS nanosheets shell as high active area pseudocapacitive material. The 3D graphene layer serves as excellent surface area to support 3D NCS/CNS; moreover, it provides excellent electrical conductivity between nickel foam current collector and the 3D NCS/NCS composite. Using these hybrid advantages the as-synthesized graphene/NCS/CNS composite electrode exhibits high areal capacitance of 15.6 F/cm2 at current density of 10 mA/cm2; excellent cycling stability of 93% after 5000 of cycles and excellent rate capability of 74.36% as current increase from 10 to 100 mA/cm2. Moreover, a prototype of asymmetric device fabricated using graphene/NCS/CNS as positive electrode and RGO as negative electrode exhibits high energy density of 23.9 Wh/kg and power density of 2460.6 W/kg at high operating current of 100 mA. Such high performance electrode material may get great application in future energy storage device.Three-dimensional (3D) core/shell structure of nickel cobalt sulfide is nano-engineered by using series of hydrothermal steps on a CVD grown graphene for supercapacitor application. This core/shell is composited of NiCo Based on energy storage mechanism supercapacitors are of two types, namely, electrical double-layer capacitors (EDLCs) in which energy storage is dominated by electrostatic charge diffusion and accumulation at the interface of the electrode and electrolyte; and pseudocapacitors in which energy storage is by reversible Faradaic reactions that occur at the electrode surface6.The advancement in use of portable electronic devices and high standards of living imposed the world to consume large amount of energy. So, to overcome this demand designing highly efficient energy storage device is an important issue. Among the energy storage devices, supercapacitors (SCs) are one of the promising devices due to their ultra-high power density, long cycling stability, wide range of operating temperature, and improved safety8. Different morphologies of ternary metal sulfides were investigated as high performance psudocapcitive materials 13. Even though ternary metal sulfides showed promising performance, practical application of ternary metal sulfides as electrode material is still limited because of two main challenges, namely, low rate performance at high current densities and short cycling stability.It is well known as the performance of SCs devices mainly depends on the properties of the materials used; and the key to achieve outstanding performance is to seek appropriate materials and designing suitable electrode architecture that support maximum efficiency of the proposed material. Recently, nanostructured nickel cobalt sulfides’ composites have attracted much research attention as pseudocapacitor electrode materials15. Recently, considerable effort has been devoted in synthesizing core/shell nanostructures for energy storage applications to increase the active area, cycling stability and rate capability of the device. For example, Kong et al.16 reported Core-shell NiCo2S4 nanostructures supported on nickel foam with capacitance value of 1.948 F/cm2 at 1 mA/cm2 and 79.36% of rate retention at 20 mA/cm2; Fu et al.17 reported Cobalt sulfide nanosheets coated on NiCo2S4 nanotube arrays as core/shell structure with highest capacitance of 4.74 F/cm2 at 5 mA/cm2 and rate capability of 47.67% at current density of 50 mA/cm2; similarly, Zhang et al.18 has reported nanoforest of hierarchical Co3O4@NiCo2O4 nanowire arrays synthesized via a facile strategy for electrochemical supercapacitors. Unfortunately, the reported structures still suffer from low capacitance, inferior rate performance and low utilization rate of active materials and low cycling performance at high currents. Even though electrode materials possessing high capacity, long cycling stability, and excellent rate capability are highly desirable, it is difficult to achieve all these objectives simultaneously from a single materials. So, hybridizing of different materials with different properties is becoming interesting research area19. Recently, hybridizing ternary metal sulfides with nanocarbon based material is becoming important research area. From nanocarbons, graphene is one of hot research area for energy storage application due to its unique properties, including high surface area, excellent conductivity and excellent mechanical and chemical stability which are desirable properties for SCs application21. In ternary metal sulfide/graphene hybrid composite, graphene serve as flexible high surface area to physically support pseudocapacitive ternary metal sulfides and excellent electron transport channels and resulted in improved capacitance, cycling stability and rate capability. Du et al.22 demonstrated facile synthesis and superior electrochemical performances of CoNi2S4/graphene nanocomposite suitable for supercapacitor electrodes with maximum specific capacitance of 2009.1 F/g at a discharge current density of 1 A/g; Wang et al.23 reported electrodeposited nickel cobalt sulfide nanosheet arrays on 3D graphene/nickel foam for high performance supercapacitors with good electrochemical performance; similarly, Patil et al.24 and Beka et al.25 reported the improvement in capacitance performance after hybridizing of ternary metal sulfides with graphene. Even though there are many works mentioned in the literature for ternary metal sulfide/graphene composite, the electrochemical performance is still in its infancy compared to the theoretical expectations. As a result, design of an integrated smart architecture ternary metal sulfide porous nanostructure that consists of 3D building blocks grown on 3D graphene is significantly desired, to use synergistic advantages of graphene and the ternary metal sulfides.In general to address these kinds of problems a wide range of nanomaterial design concepts have been developed such as decreasing the material features size to nanoscale with the structures such as core/shell, hallow particles and tubesHerein, we proposed a porous 3D nickel cobalt sulfide core/shell (NCS/CNS) nanostructure grown on 3D graphene decorated nickel foam for supercapacitor applications. Initially, the 3D graphene was grown on nickel foam using CVD process; then, using series of hydrothermal treatments 3D NCS/CNS core/shell is grown on the graphene layer. This hybrid composite uses the high surface area and excellent mechanical properties of graphene to support 3D porous NCS/CNS core/shell; moreover, the excellent conductivity of graphene leads to super electron transport channels between the collector and the active material. The 3D NCS/CNS core/shell nanocomposite contains numerous nanosheet shells which results in excellent porosity and high active area which are favorable for energy storage application. Using these synergistic effects of graphene and ternary metal sulfides the as-fabricated nanocomposite structure shows excellent specific capacitances with outstanding capacitance retention at high current densities and excellent cycling stability. To further evaluate our electrode material for practical application we assembled a prototype asymmetric device of 3D NCS/CNS core/shell/on graphene decorated nickel foam as positive electrode and RGO as negative electrode, and the assembled device receives an excellent electrochemical performance. The results obtained in this nanocomposite showed promising performance of hybrid structure of porous core/shell ternary metal sulfide and graphene for future energy storage applications.26 and labeled as GNF/NCS nanotube. Lastly, CNS porous nanosheets were grown on the NCS nanotube and the sample is labeled as GNF/NCS/CNS. Figure 27. However, the electronic and mechanical properties of graphene are strongly influenced by the thickness or number of layers of graphene present in the composite. As a result, the dimensional characterization of the as-synthesized graphene on nickel foam is crucial. So, to know the average thickness of our graphene we conducted Raman spectroscopy test on different places and almost the sample shows approximately any of the three graphs shown in Fig. 28.The fabrication process of our composite electrode is schematically represented in Fig. Figure 2S4 (JCPDS 43–1477). On the other hand, the crystal phase of the sample after the formation of shell is illustrated by using Fig. 3S2 (PDF # 24–0334). From the XRD pattern of the as-synthesized composite Co atoms cannot be directly seen, this is because the atomic radii of Co and Ni are quite similar and the partial substitution of Ni atoms by Co atoms does not change the crystal structure of Ni3S2 except change in lattice parameters29. So, substituting some of the Ni atoms by the Co atoms the composite can be indexed to a chemical composite of a form CoxNi3−xS2 where 0 < × < 3. Especially, the existence of additional peaks in the GNF/NCS/CNS core/shell composite compared to GNF/CNS sample proves the successful transformation of the nanotube structure into GNF/NCS/CNS core/shell structures.XRD diffraction pattern was used to investigate the crystal phase of as synthesized samples. In Fig. 3/2 and Co 2p1/2, respectively. The existence of 15.1 eV energy band difference between Co 2p3/2 and Co 2p1/2 spectrum confirms the existence of both Co2+ and Co3+ on the surface of the composite30. While in the core/shell composite the amount of Co is reduced and only one weak peak is observed at 781.4 eV for Co 2p3/2, suggesting the existence of only Co3+; the reduction in the cobalt amount is in agreement with the XRD analysis. Similarly, Fig. 3/2 and Ni 2p1/2, respectively. The energy band difference between Ni 2p3/2 and Ni 2p1/2 spectrum is 17.5 eV which indicates the existence of both Ni2+ and Ni3+ on the surface of the composite before and after the formation of core/shell31; while the other peaks are satellite peaks. The S 2p XPS spectrum is shown in Fig. 1/2 and S 2p3/2, respectively32. In the core/shell composite the peaks are reduced due to less number of S atoms in the shell (CoxNi3−xS2) compared to the number of S atoms in the core (Ni Co2S4). Finally, Fig. 2 hybridized carbon atoms forming honeycomb, implying existence of graphene in the composite34.The chemical states and electronic configuration of each element in the composite is studied in Fig. 35 the capacitance of pristine graphene on nickel foam is very low and we didn’t consider in this paper for comparison. The CV and GCD plots of GNF/NCS/CNS core/shell and the respective comparison with control samples are shown in Fig. 2 is shown in Fig. To measure the electrochemical performance of GNF/NCS/CNS core/shell structure we conducted comparative electrochemical tests with respect to control samples such as nickel foam (NF)/NCS/CNS core/shell and NF/NCS nanotube (without shell) structures which are grown on the pristine nickel foam (nickel foam without graphene). According to our previous report2 at current density of 10 mA/cm2. For more detail, Table 2) of GNF/NCS/CNS core/shell at different current densities. Even at high current density of 100 mA/cm2 our porous GNF/CNS core/shell shows high areal capacitance of 11.6 F/cm2. Thus, the as synthesized structure shows 74.36% capacitance retention as current increase from 10 to 100 mA/cm2. On the other hand, the NF/NCS/CNS core/shell and NF/CNS core show the highest specific capacitance of 10.9 and 8 F/cm2 at current density of 10 mA/cm2, respectively. Moreover, Fig. The specific capacitances of the as-synthesized composites are calculated using equation  and 2)..2). The 2 and the result is shown in Fig. 36.Cycling stability is one of key parameter to evaluate the performance of SCs. As a result, we conducted a comparative cycling stability test for GNF/NCS/CNS core/shell, NF/NCS/CNS core/shell and NF/NCS core electrodes by applying constant current density of 50 mA/cmESR), the high frequency semicircle diameter corresponds to the charge transfer resistance (Rct) and the more slope line in low frequency region is related to good capacitance behavior and low diffusion resistance37. The magnified scale high frequency region EIS plot is shown in Fig. ct values are 0.1, 0.155 and 0.32 Ω for GNF/NCS/CNS core/shell, NF/NCS/CNS core/shell and NF/NCS electrodes, respectively. Similarly, the approximate RESR values are 0.49, 0.5, and 0.52 Ω for GNF/NCS/CNS core/shell, NF/NCS/CNS core/shell and NF/NCS core electrodes, respectively. Moreover, the more slope line /(C−ΔV−)43, where, C+ and ΔV+ are correspondingly the mass specific capacitance and working voltage window of GNF/NCS/CNS core/shell electrode tested in the three electrode and similarly C− and ΔV− are correspondingly the mass specific capacitance and working voltage window of RGO electrode tested in three electrode. Hence, 44 mg/cm2 of RGO is required to balance 8 mg/cm2 of GNF/NCS/CNS core/shell positive electrode. Figure 43; thus, clearly shows as our asymmetric device can support a wide potential window of 1.6 V. Figure To investigate the practical application of GNF/NCS/CNS core/shell structure we assembled a prototype asymmetric electrode consisting of GNF/NCS/CNS core/shell as positive electrode and reduced graphene oxide (RGO) as the negative electrode. Asymmetric electrodes have been found to be an effective approach to meet the ever increasing demand of high energy density SCs, through providing a larger voltage window by combination of a carbon-based material as negative electrode with pseudocapacitive based materials as positive electrode. Cui 2 at current density of 15 mA/cm2. Figure 2 and illustrating excellent rate capability of 80%. The energy and power densities are calculated using a relation given in equation ,,2 at curin situ grown on graphene decorated nickel foam current collector by using stepwise hydrothermal process. The excellent capacitive performance of our GNF/NCS/CNS core/shell structure is due to the contribution of high surface area graphene which supported 3D porous NCS/CNS core/shell structure. Moreover, the 3D porous shell structure leads to increase of active area and resulted in high areal capacity. Additionally, the graphene layer sandwiched between the NCS/CNS core/shell and nickel foam current collector serves as a super high way for electron transport and leads to better rate performance. Using these synergistic advantages of graphene and NCS/CNS core/shell 3D porous structure we got superior areal capacitance of 15.6 F/cm2 at current density of 10 mA/cm2, excellent cycling stability of 93% after 5000 of cycles and excellent rate capability of 74.36% as current increase from 10 to 100 mA/cm2. This kind of hybrid structure may hold great potential for high performance energy storage devices in the future.In summary, Porous NCS/CNS core/shell structure was 2 was cleaned with deionized (DI) water and absolute ethanol (Et) and dried.All the reagents used were analytical grade without farther purification. Prior to the synthesis process, nickel foam of area 2 × 4 cm2 (100 Sccm) and CH4 (2 sccm) gas were flew into the quartz tube for 90 minutes to grow 3D graphene layer on nickel foam (GNF). Finally, the sample was cooled down rapidly to room temperature with the gas protection of H235.The pre cleaned nickel foam was placed in the quarz tube of CVD furnace and heated to 1023 °C in 80 minutes, maintained at 1023 °C for 20 minutes under argon (100 Sccm) gas atmosphere. Then, H2, 0.95 g of NiCl2, 0.72 g of urea and 60 ml of DI water were mixed by magnetic stirring until a transparent homogenous pink solution was formed and the solution was transferred to 80 ml Teflon-lined stainless steel autoclave. Then, the pre-prepared GNF was diagonal placed into the solution and the autoclave was sealed and kept in an oven at a temperature of 120 °C for 6 hour. After cooling down to room temperature, NCS nanoneedls precursor grown on GNF was washed with DI water and Et several times and dried at a temperature of 50 °C for 4 hours. Then, the resulted, GNF/NCS-precursor was hydrothermally treated in 80 ml Teflon-lined stainless steel autoclave containing a solution of 1.8 g of Na2S and 60 ml of DI water at temperatures of 180 °C for 8 hours. Then, after cooling down to room temperature and washing we get GNF/NCS nanotube.In this step, 1.855 g of CoCl2, 0.25 g of NiCl2, 1.2 g of urea and 12 ml of DI water, 12 ml Et and 12 ml of ethyl glycol were mixed by magnetic stirring until a transparent homogenous pink solution was formed and the solution was transferred to 60 ml Teflon-lined stainless steel autoclave and finally GNF/NCS was placed in the solution and it was hydrothermally treated at a temperature of 250 °C for 1 hours and after normally cooling down and washing we dried at a temperature of 50 °C for 4 hours and we got GNF/NCS/CNS core/shell.In this step, 0.5 g of CoClIt was prepared by hydrothermal reduction of GO; in detail, 120 mg of GO was immersed in 48 ml of DI water, then ultrasonicated for 5 hour at a temperature of 60 °C. Then, the solution was transferred to a 80 ml Teflon-lined stainless steel autoclave and heated at a temperature of 180 °C for 6 hrs, after cooling down to normal temperature the resulting sample is washed by vacuum filtration using DI water several times and dried at a temperature of 60 °C for 6 hr.Kα radiation (λ = 0.1542 nm). X-ray photoelectron spectroscopy (XPS) was used to study chemical state of the elements in the composite. Raman spectroscopy was carried out by HR800 with an excitation wavelength of 633 nm to study graphene on the nickel foam.To study the morphology of as synthesized sample scanning electron microscopy (SEM) Hitachi S-4800 was used. To study how atoms are arranged in crystal structures, we conducted X-ray powder diffraction (XRD) patterns on a Rigaku Max-2200 X-ray diffractometer with monochromatized Cu2 area GNF/NCS/CNS core/shell sample was sandwiched between two pieces of nickel foams and pressed at pressure of 3 MPa and used as working electrode. Moreover, we also tested the electrochemical performance of assembled prototype working device. In this case, we used GNF/NCS/CNS core/shell as positive electrode and RGO as negative electrode. The RGO negative electrode was prepared by coating a mixture of RGO, acetylene black and poly (tetrafluoroethylene) in a weight ratio of 80: 10: 10 on nickel foam. Then, the electrode was dried for 12 hour at 60 °C in a vacuum. The average areal specific capacitance and mass specific capacitance values in three electrode measurement were calculated from GCD curves, using equations (1) and (2)45:CA(F/cm2), CS(F/g), I(A), t(S), m(g), S(cm2) and ΔV(V) are specific capacitance, areal capacitance, discharge current, discharge time, mass of active material, area of active electrode and potential window for charge discharge, respectively. On the other hand, the specific capacitance of two-electrode system is calculated using the relation given by refs cell is the specific capacitance of an asymmetric cell, I is discharge current of the cell, ΔV is the operating voltage window of the cell, m’ is sum mass of active materials on both electrode  after obtaining the balanced mass of the two electrodes. Energy density and power density are key parameters to measure the practical performance of electrodes. So, the energy and power densities are calculated using the following relations:44.The electrochemical properties of the as-synthesized sample were tested by conducting cyclic Voltammetry (CV), galvanostatic charge-discharge (GCD) and electrochemical impedance spectroscopy (EIS). All electrochemical performance measurements were carried out using electrochemical workstation  using 3 M KOH aqueous electrolytes. In a three-electrode system measurement, the as-synthesized active material was used as a working electrode; Platinum plate and Ag/AgCl with saturated KCl are used as a counter and reference electrodes, respectively. A 1 × 1 cm


The use of electric fields for signalling and control in liquids is widespread, spanning bioelectric activity in cells to electrical manipulation of microstructures in lab-on-a-chip devices. However, an appropriate tool to resolve the spatio-temporal distribution of electric fields over a large dynamic range has yet to be developed. Here we present a label-free method to image local electric fields in real time and under ambient conditions. Our technique combines the unique gate-variable optical transitions of graphene with a critically coupled planar waveguide platform that enables highly sensitive detection of local electric fields with a voltage sensitivity of a few microvolts, a spatial resolution of tens of micrometres and a frequency response over tens of kilohertz. Our imaging platform enables parallel detection of electric fields over a large field of view and can be tailored to broad applications spanning lab-on-a-chip device engineering to analysis of bioelectric phenomena. Detection of electric fields, central to chemical and biological processes, has been limited to measurements of current  and secondary reporters . Here, the authors demonstrate an optical platform capable of imaging electric field dynamics with high spatio-temporal resolution. Signalling and manipulation through the control of electric field distributions is ubiquitous to both biological and physical systems. For example, intercellular electrical activity is central to the signalling and computation of excitable cells such as cardiac and neuronal cells12467810Over the past decade, there has been a concerted effort to develop new techniques to observe and analyse the dynamic field fluctuations in liquids, such as the development of multitransistorarrays1214151617101919πα∼2.3% at all frequenciesAtomically thin materials like graphene offer exceptional opportunities for electric field sensing due to their unique physical properties and intrinsic sensitivity to their environment. Over the past decade, graphene's optoelectronic properties have been studied extensively23−4. However, physical and biological applications require orders of magnitude improvement in voltage sensitivity. To increase the light-matter interaction in graphene, researchers have explored different approaches such as coupling graphene to silicon waveguides262728293132Realizing this goal requires new optical designs that optimize the detection sensitivity and parallel readout simultaneously. For example, simple transmission mode imaging of gated monolayer graphene yields a 1% change of transmitted light over a gate voltage change of 200 mV around the Pauli blocking region. This results in a voltage sensitivity of only 2 mV, accounting for a typical laser with noise levels of 10Here we demonstrate highly sensitive, high-speed optical imaging of local electric field dynamics in solutions using graphene and a critical-coupled planar waveguide. Using a custom numerical simulation, we designed the critically coupled waveguide amplified platform to obtain the so-called critical coupling condition2O5 (n=2.0856), forming a high refractive index waveguide. We use an equilateral SCHOTT SF-11 glass (SF-11) prism (n=1.743) with a 1,000 nm top layer of SiO2 (n=1.444) to evanescently couple a collimated 1.55 μm beam to and from the CAGE sensor. The reflected light is then collected by an infrared objective and imaged onto an InGaAs camera and photodiode.2 and Ta2O5/solution interfaces. Under this framework, we can describe the waveguide as a Fabry-Perot cavity with high reflection coefficients |r1| and |r2| at the two interfaces, where |r1| and |r2| deviate from unity due to the frustrated total internal reflection from the finite SiO2 thickness at the first interface and the absorption of graphene at the second interface. At resonance coupling, the total reflection R from the Fabry-Perot cavity is described by |r2| can be varied in situ by electrostatic gating of graphene. To attain the highest sensitivity to local electric fields, we gate the graphene to a specific Fermi energy that generates the largest relative change in optical reflection for a given local electric field. At the critical-coupling condition, |r1|=|r2| and the total reflection, R, equals zero, at which point all of the light is absorbed by graphene. Consequently, close to the critical coupling condition, the light-graphene interaction is strongly enhanced and the background reflection is very weak contributing to the high voltage sensitivity of CAGE imaging.The concept of waveguide critical coupling is illustrated in ibed by  . The val2 layer, the frustrated total internal reflection |r1|=98.2%. The reflection coefficient at the Ta2O5/solution |r2| is 97.5% at the charge neutral point (Vg=−0.14 V) due to graphene absorption, which leads to a total reflection of R ∼1.5%. Graphene absorption can be set subsequently by electrostatic gating through the solution. The critical coupling condition |r1|=|r2| is realized at Vg=+0.41 V (electron doped) and Vg=−0.69 V (hole doped), resulting the lowest total reflectivity, R . Further increase of carrier doping leads to a decrease of graphene absorption corresponding to |r1|<|r2| and one obtains an increase in total reflectivity. The grey dashed line in R/R caused by a change in voltage induced by the local electric field, ΔV. R/R)/ΔV as a function of the gate voltage from Vg=0.53 V. This sensitivity is over 200 times higher than that of the direct transmission configuration /ΔV=1.2% per mV reported in Vpp=100 μV with a signal to noise ratio (SNR) of 6.5. This measurement reflects optical detection capabilities of ∼15 μV or, equivalently, a change of carrier concentration of 2.3 × 108 electrons per cm2 in graphene. The noise level at 0.017% in CAGE optical detection of small electric field fluctuations is demonstrated in C is the capacitance of the graphene/electrolyte interface and R is governed by graphene conductance. For local electrical field fluctuations, the effective area and capacitance is small and the response speeds can be very high. Experimentally, we characterized the frequency response of the CAGE detector using a relatively large graphene area (200 by 400 μm), underscoring the applicability of our measurements to large fields of view. R/R value remains largely constant up to 10 kHz and decreases at higher frequencies. These measurements yield a 3 dB frequency of 13 kHz  and 150 nm Ta2O5 (waveguide layer) deposited on one face of a 1 cm equilateral SF-11 glass prism by ion-assisted deposition. The structure was designed using a custom Python simulation (2 area graphene was transferred onto the waveguide surface by poly(methyl methacrylate) (PMMA) transfer. We obtain high-quality large-area graphene with near-uniform optical absorption at the device/solution interface. The Pt (2 nm)/Au (60 nm) electrodes were deposited on the graphene to make electrical contacts. The metal electrodes were insulated with nitrocellulose lacquer to prevent water-Au chemical reactions during measurements. The device was mounted in a solution chamber printed from PR48 resin (Autodesk), which permits access to the device from both the top and bottom. All data were obtained in a saline solution  except for imaging data  was applied through a Ag/AgCl electrode in solution to set the Fermi energy of graphene and to test the optical response of CAGE detection under electrostatic gating. For the spatially resolved measurements, an external gate voltage Vg (1.1 V) was applied through the microelectrode, whose high impedance at the electrode/solution interface requires a larger applied voltage but yields the same voltage bias and critical coupling condition at the detector's graphene/solution interface. A 10 mV electrical pulse generates a 1.8 mV local field at the graphene/solution interface beneath the Pt/Ir microelectrode tip   . To thisThe data that support the findings of this study are available from the corresponding author upon request.How to cite this article: Horng, J. et al. Imaging electric field dynamics with graphene optoelectronics. Nat. Commun.7, 13704 doi: 10.1038/ncomms13704 (2016).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.Supplementary Figures, Supplementary Notes and Supplementary References.


Ultrafast and broad spectral bandwidth photodetectors are desirable attributable to their unique bandstructures. Photodetectors based on graphene have great potential due to graphene’s outstanding optical and electrical properties. However, the highest reported values of the photoresponsivity of pure graphene are less than 10 mA/W at room temperature, which significantly limits its potential applications. Here, we report a photoresponsivity of 32 A/W in pure monolayer graphene photodetectors, an improvement of over one order of magnitude for functional graphene nanostructures (<3 A/W). The high photocurrent generation in our devices can be attributed to the high sensitivity of graphene’s resistivity to a local change of the electric field induced by photo-excited carriers generated in the light-doping substrate. This dramatically increases the feasibility of using graphene for the next generation of photodetectors. As a novel two-dimensional (2D) material, graphene possesses a unique conic band structure –3 and at2) substrate. We demonstrate a higher photoresponsivity at room temperature at visible range. The photoresponsivity value reaches a maximum of 32 AW−1 at room temperature measured from biased source-drain and back-gate voltage modulation, one order of magnitude higher than the values reported in recently published works. Our work potentially reveals the importance of the effects of the doping substrate for graphene photodetectors and offers insight on how to optimize pure graphene-based photodetectors and transistors for optoelectronic and electronic practical applications.To date, work on creating graphene photodetectors with a high response speed and a broad spectral range at room temperature is still ongoing. More recent efforts concentrate on graphene functionalization techniques by creating a bandgap to enhance the photoresponsivity. However, not much work has been done on understanding and controlling the substrate effect on graphene photodetectors. Several known mechanisms contribute to photocurrent generation in biased graphene with a heavily doped silicon substrate , 24, 26.2 dielectric and confirmed with Raman spectroscopy. In order to detect the excessive photo field-effect, we used a lightly doped (p-type 10–20 ohm-cm) silicon dioxide on silicon substrate. In situ sample cleaning by the electric field were performed to clean the surface, enhance carrier mobility, and reduce intrinsic defects. After this treatment, the Dirac point of the devices were near 25 V. The photocurrent measurements were performed with a helium–neon laser with a wavelength of λ ≈ 632 nm at room temperature. Incident laser power values were fixed at 5 mW, and the frequency of the laser pulse could be adjusted from 5–5000 Hz. The laser spot diameter on the sample could be adjusted from d ≈ 1 μm to 1 mm using a microscope objective lens. The photoresponsivity data was collected with a semiconductor device analyzer . Considering the electric field enhancement attributable to the gate stack, we calculated that the absorption of the incident light in the graphene device was 2.5%. The photocurrent amplitudes shown are peak-to-background values throughout. The photoresponsivity measurements were carried out on multiple samples.Graphene photodetectors with an FET configuration were fabricated by mechanical exfoliation from bulk highly oriented pyrolytic graphite . Photolithography and e-beam evaporation of 5 and 80 nm of chromium and gold, respectively, were used to create the source and drain electrodes. The sample of monolayer graphene was identified by optical contrast on the 300 nm SiOI–V) characteristics of the graphene photodetector with back-gate bias (VG) varying from −10 to 10 V. Note that the current–voltage (I–V) with respect to Vs-d = 0 is asymmetric; this asymmetric behavior indicates that an intrinsic internal electric field  exists in the device. This plays an important role in the overall photocurrent response of graphene photodetectors, possibly as a result of both fabrication-induced defects and potential barriers between the edges of the graphene sheets. The inset in Fig. pD2/pG > 1 confirms that the sample is monolayer graphene. In addition, as the D peak is not seen in the Raman spectrum, the graphene sample must be relatively free of defects , P is the incident laser power [W], SG and SL are the area of the sample and laser spot, respectively, and λincident is the laser wavelength. The laser-induced charge carrier generation and subsequent separation at the graphene–metal interface results in photocurrent generation. When light is incident on the interface between the graphene and the electrodes, the photo-excited electron–hole pairs are generated and accelerated in opposite directions by the internal electric field, thus generating an observable photocurrent Iph = VOC/Rg .We investigated the photoresponsivity of the pure graphene photodetector device (refer to Sample A in the latter part of the paper) at source-drain voltage see Fig. . In thesVG = − 1V. As shown in Fig. Iph and photoresponsivity are strongly dependent on the size of the laser spot. The photocurrent Iph increases linearly with the laser spot size and saturates at spot size d = 1.0 mm. Such behavior indicates that the photocurrent originates from several different mechanisms [2 interface under illumination without a source-drain bias, leading to a photogating effect in the graphene FET [Iph reduces to 10 nA. This value agrees with the previously reported results of a low photoresponsivity of pure graphene of about 10 mAW−1 [To clarify the origin of the photocurrent, we investigated the photocurrent dependence on the laser spot size. This allowed us to clearly identify the photocurrent contributions from both the silicon substrate and the graphene–electrode interfaces, where internal electric fields are produced and separate the photo-generated carriers . In realchanisms , 25, 26.hene FET , 34. TheOwing to the work function mismatch between silica and silicon, the valence and conduction bands in silicon bend at the interface . In our VG stays fixed at 0 V) of sample A are shown in Fig. I is the sum of the photocurrent and dark current: I = Iph + Id. Interestingly, the photocurrent Iph increases with increasing source-drain bias voltage , but decreases for 0 > Vs − d > − 55mV and changes photocurrent sign at Vs − d < − 55mV  and gain (G) rise linearly and saturate at Vs − d = 2V. A maximum photoresponsivity of S = 0.17 AW− 1 is achieved at room temperature at a source-drain bias of Vs-d = 2000 mV, representing a 17-fold improvement over our previously reported value for our graphene photodetector at room temperature. The enhanced photoresponsivity with the addition of the substrate indicates that the lightly doped substrate significantly improves the functionality of the graphene photodetector. This result holds promise for prospective applications in a new era of high-performance optoelectronic devices.Figure V ≤ Vg ≤ 12 V) applied to the silicon back-gate at Vs − d = 5mV. With a positively biased back-gate voltage, the photocurrent Iph decreases with increasing back-gate bias voltage. In contrast, the photocurrent Iph increases with the negative back-gate biases. Figure S = 1.38 AW− 1 is obtained at a back-gate bias of VG = − 12V. These observed values are larger than those reported in literature based on pure monolayer graphene-based photodetectors at room temperature with bias voltage. The increasing photocurrent with the decreasing back-gate voltage can be attributed to the hole impact ionization effect with respect to the external electrical field. As the gate bias decreases, more holes are involved in the impact ionization process, generating more electron–hole pairs. The observed results indicate that the back-gate voltage induced doping of the graphene channel and shifted the position of the Fermi level. Interestingly, the photo-induced current and the bias back-gate voltage are linearly related, indicating that higher photoresponsivity can be readily obtained by applying a larger negative bias back-gate voltage. Furthermore, the photoresponsivity and gain can be gate-modulated, providing a convenient on-off switching control for the graphene photodetector.Moreover, the external photoresponsivity and gain can also be enhanced by applying a back-gate bias within the photocurrent generation path and by augmenting the interaction between graphene and the incident light. Figure VG) and source-drain (Vs-d) voltages. Moreover, the proposed configuration is superior to the previously reported structure, and a larger scale of the proposed device can be easily fabricated. Further device performance improvement can also be obtained through plasmonic nanostructures and waveguide-integrated configurations, providing that a method of growing high-quality graphene with high carrier mobility that is compatible with modern semiconductor technologies can be developed.In conclusion, we have demonstrated a highly effective method of increasing the sensitivity of a pure monolayer graphene photodetector by using lightly p-doped silicon dioxide on silicon as the substrate. Our pure graphene photodetector exhibits an increased photoresponsivity, one order of magnitude higher than that of similar graphene photodetectors in previous reports at room temperature. The observed phenomena in our experiments point to a clear pathway toward practical applications of pure graphene in the design of photodetectors that can be manipulated by back-gate (


PIN+]. Although overexpression of Sis1 altered the TDP-43 aggregation pattern, we did not detect physical association of Sis1 with TDP-43, suggesting the possibility of indirect effects on TDP-43 aggregation. Furthermore, overexpression of the mammalian Sis1 homologue, DNAJB1, relieves TDP-43 mediated toxicity in primary rodent cortical neurons, suggesting that Sis1 and its homologues may have neuroprotective effects in ALS.Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by selective loss of motor neurons with inclusions frequently containing the RNA/DNA binding protein TDP-43. Using a yeast model of ALS exhibiting TDP-43 dependent toxicity, we now show that TDP-43 overexpression dramatically alters cell shape and reduces ubiquitin dependent proteolysis of a reporter construct. Furthermore, we show that an excess of the Hsp40 chaperone, Sis1, reduced TDP-43’s effect on toxicity, cell shape and proteolysis. The strength of these effects was influenced by the presence of the endogenous yeast prion, [ Many neurodegenerative diseases are associated with aggregation of specific proteins. Thus we are interested in factors that influence the aggregation and how the aggregated proteins are associated with pathology. Here, we study a protein called TDP-43 that is frequently aggregated in the neurons of patients with amyotrophic lateral sclerosis (ALS). TDP-43 aggregates and is toxic when expressed in yeast, providing a useful model for ALS. Remarkably, a protein that modified TDP-43 toxicity in yeast successfully predicted a new ALS susceptibility gene in humans. We now report a new modifier of TDP-43 toxicity, Sis1. We show that expression of TDP-43 in yeast inhibits degradation of damaged protein, while overexpression of Sis1 restores degradation. Thus suggests a link between protein degradation and TDP-43 toxicity. Furthermore we show that a mammalian protein similar to Sis1 reduces TDP-43 toxicity in primary rodent neurons. This identifies the mammalian Sis1-like gene as a new ALS therapeutic target and possible susceptibility gene. In aPIN+] dependent polyQ aggregation and toxicity  dependent toxicity of an overexpressed prion-like protein Pin4C. When overexpressed in the presence of [PIN+], Pin4C formed large hyperphosphorylated aggregates that co-localized with Sis1 and reduced degradation of a ubiquitin proteasome system (UPS) reporter protein. Aggregation, hyperphosphorylation and toxicity were suppressed by overexpression of Sis1  dependent toxicity caused by overexpression of the [PIN+] prion protein, Rnq1. Here, overexpressed Sis1 appears to enhance aggregation of toxic detergent soluble Rnq1 oligomers into apparently non-toxic larger detergent resistant aggregates in the nucleus  cells [PIN+] and [pin-] yeast [PIN+] and [pin-] cells  ,49–51 wepin-] and [PIN+] cells in three strain backgrounds. The rescue appeared to be weaker in 74D-694 [PIN+] cells where TDP-43 is more toxic PIN+], bePIN+] cells reduced degradation of a UPS reporter, CG*  and [PIN+] 74D-694 cells but was more dramatic in [PIN+] cells where TDP-43 is more toxic. Furthermore, simultaneous overexpression of Sis1 and TDP-43 resulted in rapid degradation of CG*. This is consistent with the hypothesis that TDP-43 toxicity in part results from interference with the UPS system.Previous results showed that overexpression of polyQ or Pin4C in  than [pin-] 74D-694 cells and likewise has a greater effect on UPS in [PIN+] vs. [pin-] cells. By analogy, the presence vs. absence of heterologous aggregates in mammalian cells may also influence the proteostasis of disease-causing aggregates.Aggregation of TDP-43 is frequently associated with ALS and other neurological diseases –79 and oPIN+] effects toxicity and UPS inhibition is unknown. While [PIN+] dramatically enhances aggregation and toxicity of polyQ and Pin4C [PIN+] vs. [pin-] cells are indistinguishable. Nonetheless, the fact that [PIN+] prion aggregates sequester some essential chaperones including Sis1 [PIN+] cells more vulnerable to TDP-43 overexpression, e.g. the lower level of Sis1 available for the UPS in [PIN+] cells could make cells more sensitive to the inhibitory effect of TDP-43 on the UPS. Indeed, we found that overexpression of Sis1 reduces the deleterious effects of TDP-43 on cell growth, cell shape and UPS inhibition. Several genetic screens in yeast have identified modifiers of TDP-43 toxicity; however, Sis1 was not among them  and other unknown differences between strains alter the toxicity of TDP-43, repeating screens for toxicity modifiers in additional yeast strain backgrounds may reveal new modifiers.Why and how  cells  cells . In that+] cells ,38,66,71Thus, the mechanism by which Sis1 rescues cells from TDP-43 toxicity appears to be distinct from its rescue of polyQ, Pin4C or Rnq1 toxicity. Previous studies implicated a direct effect of a different HSP40 chaperone, DNAJB6 (a member of the DNAJB7 and 8 family) on TDP-43 via interactions with the TDP-43 C-terminal domain ,83. In cDBR1 knockdown)  versions of strains were obtained on YPD plates with 5 mM GuHCl and were confirmed to be [pin-] by checking for lack of aggregates after transforming with an Rnq1-GFP plasmid  and [pin-] versions of 74D-694, W303 and BY4741 were doubly transformed with GAL1 controlled TDP-43-DsRed, GAL1 controlled modifier or control empty vector plasmids listed below. Transformants were selected on SD-Leu-Ura plates. Normalized suspensions of cells taken from SD-Leu-Ura were 10X serially diluted in water and 15 μl were spotted on SD-Leu-Ura (dextrose), and 2% Gal-Leu-Ura  plates, which were photographed after 3 (dextrose) or 5  days of incubation at 30°C. We show hUpf1 because it had a bigger effect than the yeast homologue Ecm32 found in the initial screen  and [pin-] versions of 74D-694 (A), BY4741 (B) and W303 (C) carrying pGAL1-TDP-43-DsRed (p2173) or pAG415 GAL1-ccdB-DsRed (p2302) were grown on 2% galactose plates for 4 days and then imaged at the same magnification. Shown are representative fields of many fields photographed and examined.[(PDF)Click here for additional data file.S3 Figpin-] 74D-694 with pTETR-TDP-43-YFP (p2223) were selected on SD-Trp supplemented with doxycycline (10 μg/ml). Transformants were then grown in liquid SGal-Trp media with the indicated amount of doxycycline for 24 h and were examined and photographed at same magnification. The levels of TDP-43-YFP were determined by immunoblotting SDS-PAGE gels of normalized cell lysates probed with anti-TDP-43 antibodies, and anti-Pgk1 antibodies as an internal loading control.Transformants of [(PDF)Click here for additional data file.S4 FigPIN+] and [pin-] 74D-694 doubly transformed with pGAL1-TDP-43-DsRed (p2173) and pGAL-Sis1 (p1759) were grown in plasmid selective synthetic liquid media containing 2% galactose and 2% raffinose for 2 days. Cells were then crossed to [pin-] 64D-694 MATα (L2642) bearing plasmid p1185 (pCUP1-RNQ1-GFP). Diploids were selected on SD-Leu-Ura-Trp, expressed on SD-Leu-Ura-Trp supplemented with Cu++ (50 μg/ml), and examined for Rnq1-GFP dots or diffuse fluorescence, which was diagnostic for the presence or absence, respectively, of the [PIN+] prion.[(PDF)Click here for additional data file.S5 FigHSP104 does not prevent Sis1 overexpression from reducing TDP-43 toxicity. Isogenic [psi-] [pin-] versions of 74D-694 with (hsp104Δ) and without (WT) a deletion of HSP104 doubly transformed with p2042 (pGAL1-TDP-43-YFP) and p1767 (pGAL1-SIS1) or vector control p1752 (pGAL1-YFP) and p1768 (pGAL1) were selected on SD-Ura-Trp plates. Normalized suspensions of cells taken from SD-Ura-Trp were 10X serially diluted in water and 15 μl were spotted on SD-Ura-Trp (dextrose), and 2% SGal-Ura-Trp  plates which were photographed after 3 (dextrose) or 7  days of incubation at 30°C. B. Reduction of Ssa1 activity does not prevent Sis1 overexpression from reducing TDP-43 toxicity. Strain 1014 bearing a deletion of SSA2 and expressing a dominant negative allele of SSA1 (SSA1-21 ssa2Δ) and its isogenic parent strain, L3504 (WT) were doubly transformed with pGAL1-TDP-43-DsRed (p2173), pGAL-SIS1 (p1759), or vector controls (p2302 or p484). Normalized suspensions of cells taken from plasmid selective SD-Leu-Ura medium were 10X serially diluted in water and 15 μl were spotted on SD-Leu-Ura (dextrose), and 2% Gal-Leu-Ura  plates, which were photographed after 3 (dextrose) or 5  days of incubation at 30°C.A. Deletion of (PDF)Click here for additional data file.S6 Fig-5; # HR 3.18, p < 2x10-16, Cox proportional hazards analysis. Results were pooled from two independent experiments.*Rodent primary cortical neurons were dissected and transfected with plasmids encoding EGFP and TDP-43(WT)-mApple or mApple. In each case, neurons were also transfected with scrambled siRNA or siRNA targeting DNAJB1. (A) Knockdown was validated by immunocytochemistry using antibodies against DNAJB1. Scale bar, 50 μm. (B) Transfection with siRNA against DNAJB1 resulted in a 60% reduction in anti-DNAJB1 antibody reactivity  In longitudinal assays of neuronal survival, DNAJB1 knockdown enhanced the risk of death by 20% in control neurons expressing EGFP alone and in neurons overexpressing TDP43. * HR 1.20, p 0.004; ** HR 1.21, p 2.3x10(PDF)Click here for additional data file.


Patients (N = 165) with AGC treated at Peking University Cancer Hospital from October 2008 to February 2013 were retrospectively studied. Tissue samples prior to chemotherapy were assessed for PD-L1 expression using RNA in situ hybridization (an RNAscope assay) and immunohistochemistry (IHC). The correlations of PD-L1 expression to patient characteristics and clinical outcomes were statistically analyzed. PD-L1 mRNA signals were located in tumor compartments or the mesenchyme in a brown dotted or clustered pattern, and PD-L1 mRNA expression in gastric cancer was heterogeneous. PD-L1-positive expressions were observed in 33.9% (56/165) and 35.1% (46/131) patients in mRNA level and protein level, respectively. A positive relationship was found between PD-L1 mRNA and PD-L1 protein, and compared to IHC, RNAscope assay could provide an intuitional and quantitative data with potential clinical application. No statistically significant differences occurred between PD-L1 expression and clinical response to chemotherapy, or survival. However, we found that PD-L1 expression was higher in intestinal type than in diffuse type. These findings suggested that the RNAscope assay may be a promising method for patient assessment in gastric cancer clinical trials, which would be illustrated in further study.PD-L1 expression may be a predictive marker for anti-PD-1 therapeutic efficacy. No standard detection method of PD-L1 expression was available for advanced gastric cancer (AGC), which would be investigated in this study using RNA  Gastric cancer is one of the most lethal cancers, partially due to few effective therapies. Despite substantial efforts to develop better treatments, gastric cancer prognoses, especially that which is advanced is poor . To allehttp://www.clinicaltrials.gov/).Activation of the PD-1/PD-L1 signaling pathway may cause an immunosuppressive tumor microenvironment that protects tumor cells from immune surveillance and killing , 5. ThusAs a main target of anti-PD-1/PD-L1 therapy, PD-L1 expression by tumor cells may be a controversial marker for therapeutic efficacy , 9. Almoin situ hybridization in tumor FFPE tissues using an RNAscope assay is favored for its specificity and interpretative objectivity [Recently, a novel antibody-independent assay for RNA ectivity , 13. In in situ hybridization. Of these, 131 patients had samples evaluable for PD-L1 IHC. The screening diagram of eligible patients is depicted in Figure Overall, 165 patients were eligible for the study and had samples evaluable for PD-L1 RNA PD-L1 mRNA signals were located in tumor compartment or mesenchyme by brown dotted or clustered patterns, and PD-L1 mRNA expression in gastric cancer was heterogeneous Figure .No standard scoring criteria for PD-L1 expression in gastric cancer was determined, so we adopted criteria from the literature. First, PD-L1 mRNA expression occurred in 33.9% patients based on positive signals. Second, the PD-L1 mRNA positivity threshold was defined as PD-L1 expression in the mesenchyme or ≥ 1% tumor cells according to criteria from the KEYNOTE-012 trial . So 33.9P = 0.122, McNemar's test; Among all eligible patients, 131 patients had samples evaluable for PD-L1 IHC, and 46 (35.1%) patients presented PD-L1-positive expression. A positive relationship was found between PD-L1 mRNA and PD-L1 protein (P > 0.05). Positive PD-L1 mRNA expression in patients with gastroesophageal junction exceeded that of patients with non-gastroesophageal junction, but this was not statistically significant .Among all subjects, 93.3% had clinical response evaluations. Patients with partial response (PR), stable disease (SD), and progressive disease (PD) who positively expressed PD-L1 mRNA were 33.9%, 53.6%, and 7.1%, respectively, and these were not significantly different from those  who were negative for PD-L1 mRNA expression. Objective response and disease control rates of both groups were 33.9%, 87.5% and 40.4%, 77.1%, respectively  and 11.9 months (95% CI = 9.5–14.3), respectively , fluorescent in situ hybridization (FISH), and IHC for HER2 amplification and expression in breast cancer [in situ hybridization using RNAscope assay may hold promise for future research due to its specificity, reproducible, and interpretative objectivity.Alba's group compared RNA t cancer . Data sht cancer , 13. In P > 0.05). PD-L1 mRNA expression in patients with gastroesophageal junction exceeded that of patients with non-gastroesophageal junction (43.3% vs. 28.6%), although the difference was not statistically significant (P = 0.054). Also, PD-L1 expression was higher in intestinal type than in diffuse type (P = 0.010), which suggested the chronic inflammation caused by H. pylori infection or EB virus typically related to intestinal type might affect PD-L1 expression. Furthermore, PD-L1 mRNA expression in AGC as not associated with clinical response to first-line chemotherapy, PFS, or OS based on our results.PD-L1 expression has been associated with clinicopathological characteristics and prognosis in different cancers –24. We fIn conclusion, our findings suggested that the RNAscope assay may be a promising method to detect PD-L1 expression in gastric cancer patients, and might play an important role in clinical trials.A total of 165 patients with advanced gastric cancer treated in the Department of Gastroenterology of Peking University Cancer Hospital from October 2008 to February 2013 were retrospectively studied. All patients received first-line fluorouracil-based chemotherapy and had FFPE tumor sections containing at least 100 tumor cells prior to chemotherapy. Subjects provided written informed consent for their samples to be used in the research. Clinical data containing clinicopathological characteristics, clinical response and patient survival retrospectively obtained from medical records, and this study was approved by the ethics committees of Peking University Cancer Hospital.PD-L1 mRNAs in FFPE tumor samples were measured with RNAscope assay  following the manufacturer's instructions. In brief, 5 μm sections were deparaffinized, incubated with pretreatment reagents 1, 2, and 3 and room temperature for 10 min, boiled for 15 min, 40°C for 30 min, respectively. FFPE samples were hybridized with Hs-CD274-probes (ACD) at 40°C for 2 h. Hybridization signals were amplified and visualized with RNAscope 2.0 HD detection kit (Brown). Images were captured with a fluorescent microscope (Nikon ECLIPSE 80i) and the data were analyzed either by scoring or using Spot Studio software.RNAscope results were examined under a standard bright field microscope at 200–400× magnification. Positive signals showed as brown punctuate dots. PPIB and DapB were positive and negative probes, respectively, to control tissue RNA conditions and non-specific hybridization. Figure PD-L1 protein expression was determined by immunohistochemistry (IHC) assay using anti-PD-L1 antibody  according to previous procedure . BrieflyP < 0.05 with two-sided test was considered statistical significant.SPSS 16.0 software was used for statistical analysis. The association between PD-L1 expressions in mRNA level and protein level was analyzed by Kappa and McNemar's tests. The associations between PD-L1 expression and clinical characteristics were analyzed using a Chi-square test or Fisher's exact test. Kaplan-Meier survival curves and a log-rank test were used to assess patient survival.