The devices also showed excellent selectivity over other metal cations such as, Na+, K+, Mg2+, and Ca2+, suggesting the capability of working in a complex sample matrix. We further successfully demonstrated the detection of Pb2+ ions in real blood samples from children by using our aptasensors, and explored their potential applications for quantification. Our results underscore such graphene FET aptasensors for future applications on fast detection of heavy metal ions for health monitoring and disease diagnostics.Lead is a cumulative toxicant, which can induce severe health issues, especially in children’s case due to their immature nervous system. While realizing large-scale monitoring of children blood lead remains challenging by utilizing traditional methods, it is highly desirable to search for alternative techniques or novel sensing materials. Here we report a label-free and portable aptasensor based on graphene field effect transistor (FET) for effective children blood lead detection. With standard solutions of different Pb When BLL is below 75 μg/L, the reduction of children’s intelligence quotient (IQ) was reported to be associated with as wellLead (Pb) is a widely distributed contaminant due to industrial and mining activities, coal combustion, and use of leaded paints/gasoline12Some basic criteria to realize large-scale monitoring of blood Pb include good selectivity, high sensitivity, superior portability, fast response and low cost. However, due to the complicated matrix of blood, it is highly challenging to develop a detection technique, which combines all these required features. For example, some widely-used instruments in laboratories such as atomic absorption spectrometry (AAS) and inductively coupled plasma mass spectrometry (ICP-MS) have shown good selectivity and high sensitivity, but they are quite expensive and cumbersome67891011121315161718192021Graphene, the world’s thinnest conductive and elastic material, emerges as an ideal candidate. As a prototype two-dimensional (2D) material, graphene is easy to be functionalized and highly sensitive to environment, making it suitable to detect various objects, such as proteins, DNA, gas atoms and heavy metal ions192324252627282+concentrations, we obtained a detection limitation below 37.5 ng/L. Excellent selectivity to was also demonstrated over other metal cations Na+, K+, Mg2+, and Ca2+, suggesting the devices’ capability of working in a complex sample matrix. Furthermore, we successfully demonstrated the detection of Pb2+ ions in real blood samples from children by using our aptasensors, and explored their potential applications for quantification.In this work, we report a label-free and portable aptasensor based on graphene FET for effective children blood Pb detection. With Tris-HCl solutions of different Pb2+ detection, including G-quadruplex15172019212+ binding affinity and selectivity. The original 8–17 DNAzyme comprises of an enzyme strand (17E), which cleaves the RNA base on the substrate (17S) strand at the cleavage site upon binding to Pb2+. Typically, 8–17 DNAzyme based Pb2+ sensors relied on the release of cleaved 17S strand for signaling19212+ binding alone (by replacing the cleavable site rebonucleotide “A” to uncleavable deoxyribonucleotide “A” of the 8–17 DNAzyme) could give rise to significant signal changes with no need for the dissociation of the cleaved 17S strand. This could potentially improve the response speed of the 8–17 DNAzyme based Pb2+ sensors2+ binding capability of the DNAzyme. To ensure strong and specific binding of the aptamers to the graphene surface, we introduced a pyrene group to the 5′-end of 17E. This allows the aptamers to be anchored to the graphene surface of the devices through π-π stacking interactions. Moreover, this could successfully avoid the non-specific adsorption and denaturation of 8–17 DNAzyme on the graphene surface, which was reported to be a problem for many graphene based sensors2+ sensing. Such approaches have also been demonstrated to be valid to detect other heavy metal ions, such as Hg2+  2) were chosen for our studies due to higher quality, while CVD  grown graphene is believed to behave similarly. Shadow mask alignment or electron beam lithography followed by electron beam evaporation was applied to pattern the electrodes, which consist of 5 nm Ti and 50 nm Au, covered by an extra layer of 15 nm SiO2 to protect electrodes from electrochemical reactions . The atomic force microscopy (AFM) image of a single layer graphene flake is presented in 2 250 sccm, 300K) before electrical measurements were performed.The optical microscopic image of a typical graphene device is shown in back gate)-dependent resistance measurement results of a typical graphene FET device, showing the ambipolar field-effect characteristics. Both the carrier type and density of graphene can be modulated by Vback gate. In cnp (or the Dirac point in the case of monolayer graphene), indicating how the graphene flake is intrinsically doped, i.e. p/n doped when Vcnp is positive/negative or non-doped when Vcnp is equal to zero. For this particular device, Vcnp is around 18V, suggesting it is intrinsically p-doped. The feature of being sensitive to surrounding environment renders graphene an ideal sensing material for detecting charged objects. With the presence of certain charged environment, the charge doping level (or Fermi level) of graphene will be changed, resulting in a shift of Vcnp, or the resistance-Vback gate curve.We first confirmed the successful linking of pyrene to the 5′ of 17E, as supported by the UV-Vis spectrum shown in liquid gate curves within 2 minutes, which minimized the effect of solution evaporation. The pH of standard solutions was kept at neutral to avoid false response. At least two replicates were conducted for each concentration and the relative standard deviations between replicates were 2.4–15.7%, indicating good reproducibility of our measurement. The test results of standard solutions with four concentrations  are shown in cnp was around 0.2 V, indicating the p-doping nature of the device in Tris-HCl environment. After adding standard solution of 37.5 ng/L, we clearly observed left shifts of the curve, i.e. smaller Vcnp. For higher concentrations, the resistance-Vliquid gate shifted further left and Vcnp became smaller.To examine the performance of our graphene aptasensors for detection, we used 10 mg/L solution diluted with Tris-HCl to obtain standard solutions with different concentrations. The devices were immersed in the 0.5 M/L Tris-HCl solution during test to simulate human body fluid environment and build the ion liquid gate type of FETs . After adding standard solutions with different concentrations, we measured the resistance-Vcnp. For solutions with higher concentration, larger doping of electrons induces larger shift of Vcnp, which can be defined by ∆Vcnp2628cnp versus concentration  of the tested standard solutions from two representative devices  by approaching its surface, which leads to a left shift of the transfer curves or a smaller V2+, we measured the response of other commonly found ions in blood, including Na+, K+, Mg2+ and Ca2+ at a concentration of 0.1 M/L. As shown in cnp was much smaller than that from Pb2+ at a significantly lower concentration (0.5 nM/L). Therefore, our device can be used to measure real blood samples with no interference from other metal cations in blood. Such a great selectivity can be attributed to the preferential binding of 8–17 DNAzyme to Pb2+ over other metal ions. These results also suggested that functionalization of 8–17 DNAzyme to the graphene surface did not affect its Pb2+ binding affinity and selectivity.To study the selectivity of the aptasensor toward Pbcnp increased with Pb2+ concentration. In order to rule out possible background disturbance in real samples, a parallel test was performed with another non-aptamer device, with results shown in cnp on the bare graphene.To the best of our knowledge, although a few graphene FET aptasensor has been reported, none of them was used for the measurement of Pb level in real blood sample. To address this challenging issue, we tested the feasibility of our device to measure BLL in real children blood samples. Three children blood samples were collected from town of Zhuhang in Nantong , which is an industrial town with intensive wire rope production. Emission of Pb-containing smoke and dust during the wire rope manufacturing process result in elevated levels of Pb in various environmental matrix of this area. Several children living in a nearby residential district were diagnosed with hyperactivity and emotional problems owing to their elevated BLL higher than 100 μg/Lcnp. We defined these concentrations as aptasensor concentrations, which were plotted together with ICP-MS measured concentrations for comparison in We further explored the potential application of our aptasensor devices on Pb quantification in real blood samples. The results from the standard solutions . 17E and 17S was separately synthesized by Genscript Biotechnology Co.  and dissolved in deionized water (4 μM/L). Our real samples of children blood were collected from Zhuhang, Nantong, Jiangsu Province, China. The collection of blood samples and the related experiments performed in this study were approved by the center for disease control and prevention of Changshu, China, and the methods were carried out in accordance with the guidelines established by the center. The informed consent was obtained from parents of these children. A conventional electron-beam lithography process followed by standard electron-beam evaporation of metal electrodes  was used to fabricate graphene FET devices.17E and 17S were synthesized by Genscript Biotechnology Co. . The functionalization consists of two steps: firstly, we dropped the 17E solution (4 μM/L) on the devices, and waited for 30 minutes to ensure good attachment; Secondly, we dropped the 17S to form 8–17 DNAzyme on the surface of graphene. The devices were washed with deionized water to remove the aptamer residue after each step.3 in a graphite digestion system at 105 °C for 5 h, and followed by the addition of 1 mL of 30% H2O2. The digestion was continued to reach near dryness, which was diluted to 10 mL with Milli-Q water. Lead concentration in the digestion solution was measured using ICP-MS. We measured it with ICP-MS . During ICP-MS analysis, indium isotope (114In) was used as internal standard to test the instrument performance. Before tested with our biosensor, the digested samples were neutralized by NaOH (1M/L).The children blood was digested according to U.S. EPA Method 3050B through a Hot Block digestion system . Briefly, 0.5 mL of blood was initially digested with 10 mL of concentrated HNO2 layer of the wafers was used as the back gate. For the ion liquid gate type of FET devices, a gold wire was inserted into the solution as the liquid gate electrode. We applied a constant bias = 0.1 V while sweeping the back/liquid gate voltage forward at a speed of 0.01 V/s. There was a one-minute internal between each gate voltage sweep to ensure a stable state. For measurements of real samples, an initial signal offset was adjusted by adding a highly-diluted pretreated sample with concentration (~5 ng/L). The error bars arose from the upper and lower limit of multi-measurements operated in a certain concentration.NI USB-6251  was used as a voltage source and DL 1211  was used as a current amplifier. The doped silicon layer underneath the SiOHow to cite this article: Wang, C. et al. A label-free and portable graphene FET aptasensor for children blood lead detection. Sci. Rep.6, 21711; doi: 10.1038/srep21711 (2016).


There is an error in the Results subsection of the Abstract as well as third paragraph of the Results section. In both of these instances, 43.9% should be 43.1%.There is an error in the caption of


A foundational library called MORT (Molecular Objects and Relevant Templates) for the development of new software packages and tools employed in computational biology and computer-aided drug design (CADD) is described here.MORT contains several advantages compared with the other libraries. Firstly, MORT written in C++ natively supports the paradigm of object-oriented design, and thus it can be understood and extended easily. Secondly, MORT employs the relational model to represent a molecule, and it is more convenient and flexible than the traditional hierarchical model employed by many other libraries. Thirdly, a lot of functions have been included in this library, and a molecule can be manipulated easily at different levels. For example, it can parse a variety of popular molecular formats , create the topology and coordinate files for the simulations supported by AMBER, calculate the energy of a specific molecule based on the AMBER force fields, etc.http://cadd.suda.edu.cn/MORT/index.htm.We believe that MORT can be used as a foundational library for programmers to develop new programs and applications for computational biology and CADD. Source code of MORT is available at Molecular modeling techniques have been widely used in the fields of chemistry, biology, drug design, and materials science for studying molecular systems ranging from small molecules to large biological molecules and even material assemblies. A lot of molecular simulation and visualization tools or packages have been developed-9. AmberMORT is written in C++ that is a native OOP language, and a relational model and other well-designed patterns have been applied to this library, which makes it very flexible. Furthermore, many commands/functions supported by XLEAP and AmberTools have been merged into this library. Based oAn advantage of MORT is that it employs the relational model rather than the traditional hierarchical model to store all the information of a molecule. In the hierarchical model, a molecule owns some residues, and a residue owns some atoms. Therefore, a molecule does not own atoms directly. In the relational model, a molecule owns residues and atoms directly, while a residue does not own atoms, and they just have relations between each other.The hierarchical model is used by XLEAP and some other software packages . But itHowever, all these problems do not exist or can be easily fixed for the relational model employed in MORT. Firstly, the problem of iterating on atoms does not exist because a molecule owns atoms directly, and then users can perform the iteration on a molecule directly disregarding residues. Secondly, the relational model does not have any problem dealing with bonds, angles and torsions, because in the relational model a molecule directly owns bonds, angles (if any) and torsions (if any). Finally, compared with the hierarchical model, the relational model is extremely easy to be extended. Taking the strand as an example, to introduce a strand by employing the relational model, users do not need to change the old code. Strand can only appear in DNA-related program and all users need to do is to make sure that the relations between strand and residues have been created correctly.atom, bond, resd, angl, tors, oops, tor2 and ptor.The names of these object types are encoded into HASH values in order to save the time of comparing characters. Each HASH is 10 letters long and only composed of letters, digital characters and underlines. Some HASH values are predefined, and the others can be generated by using a function with a string as the input. The flowchart for the generation of a HASH value is illustrated in Figure It has been mentioned that in the relational model, atoms, bonds and residues are owned directly by a molecule, so are angles, torsions and any other possible components such as strands and improper torsions. In MORT, these objects  are called molecular objects (referred as MOs). Currently, MORT supports eight types of MOs: atom, bond, residue, angle, torsion, out-of-plane stretch , torsion-torsion interaction, and pi-torsion (the latter two are used exclusively by the AMOEBA force field). A four-character-long-code is assigned to each MO type, and they are molecule_t) owns several MOs , and each MO has its own properties and there are relations between them. In order to save the properties and relations, two variables are created. The properties are stored in components (represented by m_components), and the relations are stored in adjacencies (represented by m_adjacencys). Both of m_components and m_adjacencys are the member variables of class molecule_t. A molecule usually has several components  and has several adjacencies . Descriptions of the important classes of variables are listed in Table The basic idea of MORT is: a molecule  but in a 1D array of floating numbers. The component is implemented in this way to achieve the best space efficiency.An adjacency records the relations between two components. For each pair of connected molecular objects, there is a record in the adjacency. More specifically, it uses a 2D array to store connections.morf_t, which is the base class for all MO classes. In brief, class molecule_t is designed for the storage of data, whereas class morf_t is created for the modification, creation or deletion of objects, and it does not contain any data information.The composition of a molecule is shown in Figure molecule_t, another class (represented by class database_t) is created to store the information of a molecule: the molecule’s pointers and their corresponding names. Therefore, if a molecule needs to be modified, its corresponding pointers will be returned from the database by using the function get_mol with its name as the parameter. The structure of class database_t is shown in Figure database_t and molecule_t are both inherited from class entity_t, which is the base class for the storage of data.Except for class morf_t is created for the modification, creation or deletion of objects. In order to achieve the above goals, some functions are necessary to access the properties of MOs, and therefore several member functions have been designed as follows:As has been mentioned before, class void set_x;value_type get_x( const string& pname ) const;bool get_x;x’ in the names of these functions can be any of the following five characters: ‘i’ (for integer), ‘d’ (for double precision), ‘s’ (for string), ‘v’ (for numeric vector) and ‘a’ (for any other data type), while the value_type can be int, double, string, numvec and boost::any depending on the ‘x’. In order to accelerate the process, string can be replaced by HASH values, and these functions can be transformed into the following ones:The character ‘void set_x;value_type get_x( const long long& pid ) const;bool get_x;iter_T) or MORANGE .Two methods can be used to iterate on the MOs of a molecule: MOITER ;iter_t operator--(int);iter_t& operator + =(ptrdiff_t pdif);iter_t& operator- = (ptrdiff_t pdif);morf_t. The following two member functions of class molecule_t return the starting and ending iterators of a certain type of MO:With all these member functions implemented, MOITER works just like a pointer to class iter_t xxxx_begin ;iter_t xxxx_end;xxxx could be any of the eight 4 character ID of a molecular object type.Here, range_T). Class range_T has the following member functions:Another way to enumerate MOs is to use MORANGE  const;morf_t at(int id) const;molecule_t has the functions xxxxs that return the MORANGE of a certain type of MOs, while class morf_t has similar member functions related_xxxxs that returns the MORANGE of related MOs.MORANGE in a sense works just like an array of MOs. Class molecule_t), and are composed of MOs . Molecules are stored in database (represented by class database_t). Both of molecule_t and database_t are inherited from class entity_t. Class morf_t and its child classes are used to modify, add and delete MOs. The functions in MORT can be roughly divided into two categories: object-related and property-related.A lot of functions supported by Antechamber and XLEAP have been developed in MORT, and therefore based on MORT it is very easy for users to develop new applications for computational biology and CADD. As shown in Figure Functions in this category can handle the MOs defined in MORT. These functions have various usages, and they are distributed in different directories.OBJFUN: a lot of functions are defined in this directory, and they can be used to modify objects. For example, the function fixbond can be used to fix the bond order of a molecule based on several rules, such as hard rule, length rule and conjugation rule, as illustrated in our previous work" may represent a carbon isotope. After the judgment, the corresponding functions will be used to analyze the character: parse_charge can be used to get the charge information from "+" and "-"; parse_ring can be used to recognize the digit outside a bracket as a starting/ending point of a ring; parse_weight can be used to parse the element’s weight that is listed in a bracket; parse_alpha can be used to interpret the element’s type, etc. Finally, all the information will be recorded into a molecule.(3). d SMARTS. SMILES CAPBOX: functions in this directory are mainly designed to add the solvent environment to an object. Four functions were designed to build different kinds of solvation environments: solvatecap can be used to add a solvate cap around the specified position of a solute; solvatebox can be used to add a solvate box to the solute in a cuboid way; solvateoct can be used to add the solvate box to a solute in a truncated octahedron way, which can reduce the number of the added solvent molecules; solvateshl can be used to add a solvate shell around the whole solute. The algorithms of these four functions are very similar. The process of solvateshl is illustrated in Figure (4). addions can be used to add positive/negative ions to the whole molecule.Apart from the functions mentioned above, the function ATMASK: sometimes, users may want to get the selected partition of a molecule, such as the atoms within 5Å from the 4th atom of a molecule. To achieve this goal, the function mask_atom was designed. This function can be employed to the specified atoms and residues. The argument of this function is a little complicated, and it is composed of some figures and symbols. For example, ":1-10" means to get the residues 1 to 10 from a molecule; "@4 < @5" means to return all the atoms within 5Å from the atom 4.(5). atom_node will then be invoked, and the following letters or numbers will be parsed as a MO in a specified molecule (here "4" represents the 4th atom). The symbol "<" means to get the atoms/residues in a certain distance, and then it will invoke the function parse of class dist_node to interpret the next section. The previous MO will be regarded as a core. The second "@" indicates that the queried objects are atoms, and "5" equals to the threshold of the distance. Finally, the function match checks each atom in the molecule that satisfies the condition, and returns the qualified ones.Following is the steps to interpret "@4 < @5". The first "@" indicates that this is something about atoms ("@" represents atoms and ":" stands for residues). The functions of class FORMAT: functions defined here can be used to handle different kinds of formatted files. For example, the function read_sdf can be used to parse the molecule from a MOL/SDF formatted file, and the function load_mdb can be used to load the molecules from a database file, which contains a lot of molecules with the MOL2, MOL/SDF or OFF format. Once a molecule has been loaded, it will be stored in a database (represented by class database_t) with its name as the identity.(6). The functions in this category can add, modify, save and delete different properties of a molecule, and all these functions are distributed in their corresponding directories.ENEFRC: functions in this directory are energy-related. The function eval_bond, eval_angl or eval_tors can be used to calculate the energy of bonds, angles or torsions. The non-bond energy can be calculated in two ways. The function get_dir can be used to calculate the energy based on periodic boundary condition (PBC) by using Ewald summation[nonbond_egb can be used to calculate the polar contribution of desolvation for a non-periodic model by using the Generalized Born (GB) model[(1). ummation,16. And Ke,C and NA are electrostatic constant, Coulomb constant and Avogadro’s contant, respectively, ε is the dielectric constant of water (78.5), INVCHG2 is a constant value of 332.05 kcal/mol, rij is the distance between two atoms, αi and αj are equal to the Born radii of atoms i and j. More detailed descriptions of the energy-related functions can be found in Additional filewhere AMBFMT: In this part of MORT, some functions were designed to read the AMBER and AMOEBA force fields and save the properties of a molecule into topology files. For example, write_amber_prmtop can be used to generate two files: topology  and coordinate files (space information), which can be used as the input files for MD simulations. read_frc/read_amoeba_frc can be used to get the AMBER/AMOEBA force field parameters from an AMBER/AMOEBA parameter file and then store them into molecules.(2). For the better use of MORT, several commands have been defined in directory "plugin", which makes it very convenient to develop one serviceable application with just a few or sometimes just one command for the developers.PLUGIN: commands in this directory can be regarded as the MORT’s interface, and users can call these commands to accomplish many kinds of tasks with less effort than using functions. They can help the developers do more work in less time.(3). In each command class, apart from its constructor and destructor, one function is necessary:virtual bool exec;command_i, which is the base class of all the command classes. exec performs variously in command_i’s child classes, reflecting the polymorphism of C++ language. Once one command class is declared, exec should be used to execute the corresponding commands at the end.This function is a virtual function inherited from class source can be used to interpret the files displayed in directory "dat/cmd", which contains the commands used for loading the force field parameters for the preparation of other operations ; merge can be used to put the objects listed in its arguments together; solvate can be used to add the solvents around a molecule; moloper can be used to add the missing information to a molecule (mentioned in directory OBJFUN part), etc. These commands are all defined in their own classes, and users need to declare the class first before doing any operation. The details of the important functions and classes in MORT and the related information of important commands can be found in Additional fileA lot of commands have be defined, for example, fixbond, addHs and other functions can work together to add the missing information.MORT can be used as a foundational library to develop new programs or software packages for computational biology and CADD. Usually, based on MORT, only a few codes are needed to solve a problem. For example, if users want to calculate the energy of a protein, the energy-related functions can be used conveniently. If the information of a molecule is incomplete, test directory.MORT can serve as the core of one program, what users need to do is to package this core with some necessary codes. They can be compiled as a standalone program to solve different kinds of problems. Figure A C++ based library MORT is developed as a new foundation library for computational biology and CADD. This new library has many advantages, especially for its data structure and powerful functions. By employing the relational model instead of the hierarchical model to store data, less time is taken while iterating on the atoms and it solves the annoying problems such as determining the belongings of inter-residue bonds. In this model, all bonds and atoms are independent and extra entities are created to store the connection information and other relations between these entities. A lot of functions have been developed in this library, and Figure setpchg can be used to call the standalone program in Antechamber[parmchk can be used to call Parmchk to add missing force field parameters. With all these commands implemented, it is easy to create the topology file for a protein-inhibitor complex inside MORT without calling any outside programs.Moreover, many commands in AmberTools have been integrated into MORT. For example, echamber to assighttp://cadd.suda.edu.cn/MORT/index.htm, and it can be compiled into a static library on Linux platform. It’s written in C++ and the boost library (version 1.46.1 or newer) is needed.MORT is available atThe authors declare that they have no competing interests.TH, QZ and WZ instigated the project, designed and implemented the algorithms and developed the MORT library. TH, QZ, YL, JW and JZ drafted the manuscript. All authors read and approved the final manuscript.The code organization, naming rules, and the detailed descriptions of the important classes and functions of MORT.Click here for fileSource code of MORT.Click here for file


TDP-43 is the major disease-associated protein involved in the pathogenesis and progression of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin-positive inclusions linked to TDP-43 pathology (FTLD-TDP). Abnormal phosphorylation, truncation and cytoplasmic mis-localization are known to be the characteristics for the aggregated forms of TDP-43, and gain of toxic abnormal TDP-43 or loss of function of physiological TDP-43 have been suggested as the cause of neurodegeneration. However, most of the post-translational modifications or truncation sites in the abnormal TDP-43 in brains of patients remain to be identified by protein chemical analysis. In this study, we carried out a highly sensitive liquid chromatography-mass spectrometry analysis of Sarkosyl-insoluble pathological TDP-43 from brains of ALS patients and identified several novel phosphorylation sites, deamidation sites, and cleavage sites. Almost all modifications were localized in the Gly-rich C-terminal half. Most of the cleavage sites identified in this study are novel and are located in N-terminal half, suggesting that these sites may be more accessible to proteolytic enzymes. The data obtained in this study provide a foundation for the molecular mechanisms of TDP-43 aggregation and ALS pathogenesis. TARDBP gene, is a highly conserved, ubiquitously expressed nuclear protein. It has conserved RNA recognition motifs (RRM1/RRM2) flanked on either side by N-terminal and glycine-rich C-terminal domains that mediate protein-protein interactions345678910TARDBP gene causes early embryonic lethality12Transactivation response (TAR) DNA-binding protein 43 (TDP-43), encoded by the otifs RRM/RRM2 flaTDP-43 is the major disease-associated protein involved in the pathogenesis of amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD) linked to TDP-43 pathology (FTLD-TDP)1416171819212223TDP-43 is predominantly localized in the nucleus, but under pathological conditions it is translocated to the cytosol14152627282914Immunoblotting of the Sarkosyl-insoluble fractions from FTLD and ALS cases using phosphospecific antibodies clearly demonstrated that hyperphosphorylated full-length TDP-43 of 45 kDa, smearing substances, and fragments at 18–25 and 35kDa are the major species of TDP-43 26312627TDP-43 phosphorylation, cleavage and cytoplasmic mislocalization are all associated with TDP-43 aggregation. However it remains to be clarified whether these modifications are the cause of the disease. Recent reports on post-translational modifications, including fragmentation, were based on studies of cellular or animal models overexpressing TDP-43 or its derivatives343536Immunoblot analysis revealed that abundant abnormal TDP-43 was recovered in the Sarkosyl-insoluble fractions of brains from both ALS cases as shown in After SDS-PAGE, the gel portions corresponding to these bands were cut out as shown in We identified at least four intrinsically cleaved TDP-43 peptides in the pathological TDP-43, i.e., three N-terminal peptides  and causes mislocalization of TDP-43 in motor neuronsAccumulation of filamentous inclusions composed of abnormally phosphorylated full-length TDP-43 (45 kDa) and its fragments (35 and 17–27 kDa) is a defining feature of TDP-43 proteinopathies3034However, no such TDP-43 peptides cleaved by caspase or calpain reported were not detected in the pathological TDP-43 in the brains of patients in this study. On the other hand, uncleaved peptides by caspase or calpain were detected as the major molecules in the digests. Furthermore, several novel intrinsically cleaved TDP-43 peptides were identified in the pathological TDP-43, i.e., three and five N-terminal peptides   and two Cellular expression of TDP-43 is tightly regulated at the transcriptional and post-translational levels. Under pathological conditions, however, formation of TDP-43 aggregates within the cell nucleus or cytoplasm results in reduced free nuclear TDP-43, and therefore the TDP-43 binding-receptor sensor detects a fall in protein levels and responds with increased TDP-43 production; the produced TDP-43 is then sequestered by TDP-43 aggregatesin vitro and in vivo4344It has been recently reported that C9ORF72 repeat expansions in mice cause TDP-43 pathology. In these mice, insoluble phosphorylated TDP-43 was predominantly present in monomeric form and high-molecular-weight or truncated pTDP-43 species were not detectedWe identified some intrinsically cleaved TDP-43 peptides at the N-terminus and C-terminus of Sarkosyl-insoluble TDP-43 in this study. These cleavage sites were neither the caspase cleavage sites nor the calpain cleavage sites of TDP-43 as reported previously46in vitro experimentIn our previous studies, we identified phosphorylation sites by immunochemical methodsIt has known that non-phosphorylated intact TDP-43 forms fibrous aggregateFurthermore, it seems reasonable to speculate that heavily phosphorylated peptides may not be detected because of low recovery from the column and low ionization efficiency in mass analysis. S409/S410-phosphorylated TDP-43 was immunochemically included in almost all fractions as shown in 2627Indeed, there were individual differences in other modifications, as shown in Interestingly, inclusions within the brain of ALS and FTLD-TDP patients are readily labelled with antibodies that recognize the C-terminus of TDP-43, but not with N-terminal TDP-43 antibodies. In contrast, spinal cord inclusions are labelled with both N- and C-terminal TDP-43 antibodies, suggesting that they are composed of full-length TDP-43 It has already reported that a Gly-rich region (287–322), which contains multiple glycine repeats, may contribute significantly to fiber formation as well as aggregation propensity5033Sarkosyl-insoluble abnormal TDP-43 was prepared from brains of two ALS cases with abundant and widespread TDP-43 pathologies. Brain samples (0.5 g) from patients with ALS  and from vascular dementia  were homogenized in 10 ml of homogenization buffer . Sarkosyl was added to the homogenates , which were then incubated for 30 min at 37 °C and centrifuged at 20,000 g for 10 min at 25 °C. The supernatants were centrifuged at 100,000 g for 20 min at 25 °C. The pellets were further washed with sterile saline and centrifuged at 100,000 g for 20 min. The resulting pellets were used as Sarkosyl-insoluble fraction. This study was approved by the research ethics committee of Tokyo Metropolitan Institute of Medical Science (number: 15–5), and carried out in accordance with the approved guidelines. Informed consent about the brain donation was obtained from all subjects.The Sarkosyl-insoluble, SDS/urea-soluble fractions were separated on 4 ~ 20% polyacrylamide gradient gels by SDS-PAGE and immunochemically detected as described-123  was used at a flow rate of 300 nl / min with a 2–80% linear gradient of acetonitrile for 60 min. Eluted peptides were directly detected with an ion trap mass spectrometer, Velos Pro  at a spray voltage of 1.9 kV and a collision energy of 35%. The mass acquisition method consisted of one full MS survey scan followed by an MS/MS scan of the most abundant precursor ions from the survey scan. Dynamic exclusion for the MS/MS was set to 30 seconds. An MS scan range of 400–2000 m/z was employed in the positive ion mode, followed by data-dependent MS/MS using the CID or HCD operating mode on the top 10 ions in order of abundance. The data were analyzed with Proteome Discoverer , Mascot software  and Scaffold software . Swiss prot and GenBank databases were used.The sample was resuspended in 0.1% formic acid containing 2% acetonitrile and introduced into a nano-flow HPLC system, DiNa fitted with an automatic sampler . A packed nano-capillary column NTCC-360/75-3How to cite this article: Kametani, F. et al. Mass spectrometric analysis of accumulated TDP-43 in amyotrophic lateral sclerosis brains. Sci. Rep.6, 23281; doi: 10.1038/srep23281 (2016).


The graphene FETs fabricated on poly (PEN) are found to have a lower density of charging sites than those on SiO2/Si substrate, mainly due to reduced interfacial interaction between the graphene and the PEN. This method can be an indispensable means to improve the stability of devices using a graphene as it provides an accurate and quantitative way to define the quality of graphene after the device fabrication.Defects of graphene are the most important concern for the successful applications of graphene since they affect device performance significantly. However, once the graphene is integrated in the device structures, the quality of graphene and surrounding environment could only be assessed using indirect information such as hysteresis, mobility and drive current. Here we develop a discharge current analysis method to measure the quality of graphene integrated in a field effect transistor structure by analyzing the discharge current and examine its validity using various device structures. The density of charging sites affecting the performance of graphene field effect transistor obtained using the discharge current analysis method was on the order of 10 The electrical and physical properties of graphene have been extensively investigated for diverse applications such as electronic switch, sensors, transparent electrodes, fuel cells, and catalysts1234568910111314For example, the conductivity of graphene is found to be greatly affected by many factors such as metallic residues originating from the growth processes, substrate materials, capping dielectrics, device fabrication processes such as patterning and cleaning, initial defect density, contact metals, operation ambient, temperature, and more. In addition, the operation history-dependent device drifts such as hysteresis, charge trapping, and interfacial reactions also affect the characteristics of a device using a graphene.3 type defects can be measured using the shift in the relative position and width of the G-peak, D-peak, and 2D-peak of the Raman spectrum. Various TEM and STM analyses of grain size, defects, and contaminants have been reported in the literature19Since the defects in graphene and the influences of the environmental factors play a critical role in device-level operation1718While Raman spectroscopy, AFM, TEM, and STM are useful means in the analysis of the initial quality of graphene, these methods are not easy to use when examining graphene integrated in a device structure2122Thus, the characteristics of graphene integrated in a device structure have been primarily investigated indirectly using the electrical characteristics of devices such as current-voltage (I-V) or capacitance-voltage (C-V) characteristics222327In this work, a new electrical characterization method to obtain quantitative information on defects and other environmental factors representing the quality of graphene FETs has been proposed and its validity has been examined using devices fabricated on a large area monolayer graphene sheets with different levels of physical defect density (see Methods). This method, called the discharge current analysis (DCA) method, modulates the carrier concentration of graphene periodically using an external pulse bias and translates the frequency dependence of the charges discharged from the graphene channel into a density of charging sites. The validity of this DCA method is examined by correlating the electrically measured density of charging sites with the initial physical defect density of graphene measured with Raman spectroscopy.For silicon MOSFETs, a charge pumping (CP) method has been used to accurately analyze the interfacial defect density, which affects the device performance. The CP method fills up defect sites at the silicon-dielectric interface using minority carriers supplied from the source and drain at an inversion state, as shown in δI, highlighted in red in oI, does not change as a function of frequency because it is proportional to the total width of the pulse peak as shown in δI, discharged from the charging sites in each cycle increases as a function of frequency. Thus, the discharge current can be approximately represented using the following equation, cI is the measured discharge current, oI is the current due to the charges accumulated in the graphene itself, k is a frequency-dependent loss factor to account for the charge loss to the source side due to asymmetric metal contacts and other factors, δI is the additional discharge current from charging sites, and f is the measurement frequency.The baseline current, cI, is linearly proportional to the frequency in the 10 KHz to 100 KHz range. As mentioned above, the tail current, δI, consists of charges released from defects sites or other surface states. The slope slightly increases above 100 KHz and then saturates at a higher frequency.Typical discharge current measured at 10 KHz to 1 MHz from CVD-grown graphene FETs is shown in cI curve, the charge trapping and detrapping mechanism in the graphene should be explained first. Recently, our group suggested that the hysteresis of a graphene I-V curve is primarily caused by two representative mechanisms, tunneling and a surface redox reaction16To understand the frequency response of the cI primarily monitors the discharge current due to the tunneling component, and it is proportional to the initial defect density as shown below. At this frequency range, cI linearly increases in proportion to the frequency. Thus, the slope of the linear portion can be used to extract the density of charging sites. On the other hand, at a frequency above 100 KHz, charge traps that cannot release charges within 5 μs start to continue holding the charges and those traps no longer contribute to cI. As a result, the slope of cI increases and eventually becomes saturated as shown in steps 5 and 6 of In the 10 KHz to 100 KHz range, c-f curves for four representative devices with different initial defect densities. All four curves show two regions of different slopes as a function of frequency at 20 to 100 KHz and 100 to 150 KHz, respectively. The cI curves are saturated at ~200 KHz and above. The linear slope at 20 to 100 KHz is attributed to the charge exchange between the charge sites and the graphene through the tunneling mechanism explained above. To investigate the influence of initial defect density (defined as I(D)/I(G) ratio) of graphene, we conducted Raman spectroscopy on each graphene channel after the graphene channel patterning /I(G) as shown in δI value representing the density of charging sites originating from both the graphene bulk channel and various charging sites near the graphene can be used to extract the density of graphene defect sites even after device fabrication.tterning , and theoI, due to the Dirac point shift during the measurement. Steps 5 and 6 of oI. The second slope showed a strong temperature dependence, indicating that it is related to the thermally activated mechanism such as chemical reaction./I(G) /I(G) . Nchargic-f measurement method can be instrumental in analyzing graphene quality, especially after device fabrication, because this method can be used to compare the impact of processes on the same graphene even without calibration.There is appreciable data scattering in c-f) curve is used to analyze the density of the charging sites while the electron-hole pair recombination at the interfacial charging sites is measured at a fixed frequency in the charge pumping method.The DCA method may look similar to the charge pumping method, which measures the interface state density of silicon MOSFETs, but the measurement principle is quite different31c-f curves measured in bottom gate graphene FETs with/without a 30 nm Al2O3 passivation layer. Since the Al2O3 passivation decreases contamination from the graphene surface, the density of the charging sites decreased to 2.47 × 1014/cm3 from 3.19 × 1014/cm3 after passivation. A more prominent slope increase before the passivation in the 100 to 180 KHz region indicates that the graphene surface is more prone to the water redox-induced charge generation, as expected2536372/silicon substrate with a poly (PEN) substrate  process was transferred to 90 nm SiO2O3 using atomic layer deposition (ALD) process at 130°C and annealed in N2 ambient at 200°C for 30 min to minimize the influence of ambient.For the device fabrication, 100 nm Au electrode patterns were formed on a graphene sheet using i-line contact photolithography and a lift off process. Then, graphene channels between source and drain electrode were patterned using i-line photolithography and oxygen plasma ashing process . The surface of graphene channel was passivated with 30 nm AlAfter the graphene channel patterning, the quality of graphene channel defined as I(D)/I(G) ratio was measured for all devices using Raman spectroscopy . 2 gate dielectric, using a pulse generator (Agilent 81110) as shown in For discharge current analysis, square pulse biases in 10 KHz to 500 KHz range were applied to the substrate, which works as a bottom gate electrode with 90 nm SiOU.J. performed electrical measurement and data analysis and worth the main manuscript with M.-H.H. and B.H.L., J.J.K. and H.J.H. performed the electrical measurement of devices and data analysis. Y.G.L. programed lab view software to collect the data presented in Fig. 2 and Fig. 3. C.G.K. and S.L. fabricated graphene FET on oxide and PEN substrate used in Fig. 3 respectively. S.K.L. developed the graphene process and fabricated the graphene sheet used in this work. M.-H.H. and B.H.L. wrote the main manuscript and jointly developed a model for data analysis. All authors reviewed the manuscript.Supplemetal data


We find that the substrate is, in fact, of great importance in the defect formation process, as the defects in graphene are mostly generated through an indirect process by the sputtered atoms from the substrate.Although ion beam technology has frequently been used for introducing defects in graphene, the associated key mechanism of the defect formation under ion irradiation is still largely unclear. We report a systematic study of the ion irradiation experiments on SiO Graphene has recently attracted great attention for a variety of novel applications, such as in nanoelectronics, gas detectors, solar cells, and DNA sequencing123468In ion-matter interaction theory, an energetic ion transfers its energy to the target system by both the elastic collisions with the target nuclei and the inelastic interactions with the target electrons (e.g. causing electronic excitations and/or ionization). In most of the previous reports112 substrate. Contrarily, Mathew et al.2 substrate under 35 keV C ion irradiation had negligible effect on the graphene. As described above, many apparently contradictory claims have been made, and the key mechanism of the defect formation under ion irradiation is still a mystery.On the other hand, the effect from the substrate underneath the graphene was also found playing an important role in the defect formation process. Compagnini et al.Y) in graphene under energetic ion irradiation. A variety of ion species with a large range of the ion energy were used. We further performed MD simulations to compare the experimental results in order to understand the underlying mechanism. We found that the defect formation only depends on the nuclear collision until eS reaches a threshold of ~ 2.2 keV/nm, above which the electronic energy loss becomes important; moreover, the majority of the defects are actually created by the sputtered atoms from the substrate rather than the direct ion-graphene collisions. Therefore, the interaction between the energetic ions and the substrate should be paid great attention to, if one would like to use the ion beam technique for engineering graphene.In this work, we report a systematic experimental study of the defect yield  monotonously increases with nS, following almost a linear relation in the log-log plot. But no such positive correlation was found with respect to eS, although the electronic energy loss is in general the major energy loss pathway for MeV heavy ions . Nevertheless, the indirect process also includes the defects generated by the recoiled incident ions16; however, they are very rare as found in our simulation, so they are safely ignored in the present discussion. Note that, by this indirect process, the defects can be generated fairly far from the incident spots. Statistically, we choose the defect-to-incident-spot distance of 5 Å as the dividing point to separately count for the defect yields by the direct process (dirY) and the indirect process (indY). As the results shown in nS used in this work: the defects generated by the direct process are consistently at least one order of magnitude less.As for the nuclear collisions, one would expect that the defects are generated by direct collision process (direct process) where incident ions directly knock out the graphene carbon atoms and leave the vacancy-type defects in graphene. However, by carefully analyzing the simulation results, we found that the majority of the defect sites were laterally quite far (a few nanometers) from the incident spots of the impacting ions, thus they were unlikely generated by the direct process. In fact, they were generated by the sputtered atoms from the SiOdT) for a carbon atom to be knocked out from a freestanding graphene sheet is about 25 eV using the AIREBO potential2, the displacement energy depends on the direction of the atoms being knocked out. We previously show that2 substrate, while it still remains around 25 eV for being knocked out into the vacuum. The former situation corresponds to the direct nuclear collision process with the impacting ions, thus the associated defect generation is greatly suppressed due to the high dT. The latter situation (towards vacuum) corresponds to the collisions initiated by the sputtered atoms from the substrate. Generally, the sputtered atoms carry energies spreading in a fairly wide range from several eV to several tens of eV following Thompson distribution2 as an example  is fairly close to 1, so the defect formation cross-section follows a simple linear relationship with respect to the nuclear stopping power. This simple linear relationship suggests that the simple binary collision model can still be used to describe the defect formation by the direct process with only a slight modification on the displacement energy (resulting in the decreased prefactor). On the other hand, for the indirect process, the defect formation is rather complicated; the cross-section depends on many factors, such as the interaction between the impacting ions and the substrate, the sputtering yield of the substrate, the types of the sputtered species, and their kinetic energy distributions. Nevertheless, the defect yield still follows reasonably well with the power law of nSends. The fitted parameters are practically useful, as they represent some universal features of the defect formation process, and can be used as a first-hand estimation for choosing the ion irradiation conditions in future.Based on the above analysis, we separately fit the direct and indirect defect yields by the power laws with respect to the nuclear stopping power in graphene  and 6 MeV Si (3.01 keV/nm) ions used in this work. On the other hand, we noticed that the substrate SiO2 has a considerably lower thermal conductance of only 1.44 W/mK2. In fact, we previously showed that the sputtering yield of bare SiO2 started to increase when eS of the incident ions exceeds 2.2 keV/nm30. Thus, the increased defect formation was most likely to be formed by an indirect process in which the sputtering yield of SiO2 was enhanced by the high electronically lost energy of the incident ions. Accordingly, for this graphene-on-SiO2 system, the threshold of eS should be determined by the substrate, i.e., 2.2 keV/nm. In addition, it is worth mentioning that, via the strong electronic excitations, part of the sputtered particles might be ionized, like oxygen radicals, which are known to chemically react with graphene . As reported before31eSshould be largely reduced. In fact, we found that, the experimental defect yield of graphene on copper was much reduced compared to that of SiO2 supported graphene (see 12 ions/cm2).For the ions in Group B, the experimental defect yields were clearly higher than the MD simulation results, as in which only nuclear collisions were considered. The experimental yields for 3 MeV and 6 MeV Si ions were roughly two times higher than the simulated counterparts, and the discrepancy became much larger when 250 MeV Sn ions were used (at least 10 times higher). This is because the electronic energy losses of these ions are so high see  that thehene see  online, 2-supported graphene were further analyzed from the MD simulation results, and compared with the suspended graphene. Taking 3 MeV Si ion irradiation as an example, typical simulation snapshots are shown in 2-supported graphene, whereas, in the suspended graphene, large complex defects (pores) were the majority. Histograms comparing the size distributions of the defects are shown in 8, and molecular adsorptionThe defect patterns in the SiO2-supported graphene by systematic ion irradiation experiments and MD simulations. Our results clearly demonstrated the great importance of the substrate during the defect formation process. The direct nuclear collision process between the impacting ions and graphene carbon atoms can also generate defects, but in much less yield (at least one order of magnitude less) than the indirect process. Simple power-law fittings of the defect yields with respect to the nuclear energy losses were given for both the direct and the indirect processes, which can be, in future, used as a first-hand estimation for choosing the ion irradiation conditions. As for the electronic energy loss of the impacting ions, this portion of energy usually does not generate defects in graphene, unless the electronic energy loss of the ions is above a threshold of 2.2 keV/nm. We further reasoned that this threshold is actually set by the SiO2 substrate, as this value is the turning point for the sputtering yield of bare SiO2 to increase, which again high lights the importance of the substrate effect. In addition, our MD simulation also showed that the small-size point defects are preferentially formed in the supported graphene. Lastly, we believe that, by choosing the right substrate along with the right irradiation conditions, we can create defects in graphene with a highly controllable manner by the ion beam technique.In summary, we have carefully investigated the mechanism of the ion-irradiation induced defect formation in the SiO2 substrate were used in the ion irradiation experiments. The monolayer graphene was grown by CVD on a copper foil, and was then transferred onto a SiO2(300nm)/Si substrate by standard transfer processY) in graphene, i.e., the average number of the defect sites produced by one single incident ion. This defect yield was derived, based on a phenomenological model19D band and the G band (D/IGI) in the Raman spectrum of the irradiated graphene. The G band (~1580 cm−1) is associated with the in-plane stretching motions of the sp2-bonded carbon atoms, whereas the D band (~1345 cm−1) corresponds to the defects in graphene36Samples of monolayer graphene on SiO2 substrate and graphene, were listed in 11 to 3 × 1014 ions/cm2.C, Si ions irradiations were carried out on the 2 × 1.7 MV tandem accelerator at Peking university in Beijing, and Xe ions irradiations were carried out on the 320 kV ECR platform at IMP (Lanzhou). Ion energies, together with their stopping powers in SiO2 slab  was 196.52 × 204.23 Å2, and the thickness was chosen to be around 60 Å. The adaptive intermolecular reactive empirical bond order (AIREBO) potential function2 slab was assumed to be van der Waals (vdW) type, and was modeled by the Lennard-Jones potential with the parameters taken from Ref. For the simulation of the ion-solid interaction process, we used MD simulation to simulate this process in order to carefully investigate the mechanism of the defect formation in graphene under ion irradiation. In our MD simulation, the lateral size of the graphene/SiOW.S.L. did the irradiations and Raman spectrum analysis of all the samples. W.S.L., X.T.Z., and S.J.Z did the MD simulations and defect statistics. W.S.L., X.W.W., H.L.D. and J.M.X. have written the paper. All authors have reviewed the manuscript.Supporting information


Here, we study large areaMoS2-graphene-heterostructures formed by direct transfer ofchemical-vapor deposited MoS2 layer onto epitaxial graphene/SiC. We showthat via a direct transfer, which minimizes interface contamination, we can obtainhigh quality and homogeneous van der Waals heterostructures. Angle-resolvedphotoemission spectroscopy (ARPES) measurements combined with Density FunctionalTheory (DFT) calculations show that the transition from indirect to direct bandgapin monolayer MoS2 is maintained in these heterostructures due to the weakvan der Waals interaction with epitaxial graphene. A downshift of the Raman 2D bandof the graphene, an up shift of the A1g peak of MoS2 and asignificant photoluminescence quenching are observed for both monolayer and bilayerMoS2 as a result of charge transfer from MoS2 to epitaxialgraphene under illumination. Our work provides a possible route to modify the thinfilm TDMCs photoluminescence properties via substrate engineering for future devicedesign.Two-dimensional layered transition metal dichalcogenides (TMDCs) show great potentialfor optoelectronic devices due to their electronic and optical properties. Ametal-semiconductor interface, as epitaxial graphene - molybdenum disulfide(MoS Recently Han et al.developed a novel seeded CVD patterned growth method to obtain highly crystallineMoS2 flakes on oxidized silicon substrate2 grown by this approach has high crystallinity, with large flakes(between 20–100 μm) and electrical and opticalproperties comparable to exfoliated material. These CVD-grown flakes are suitable fortransfer onto epitaxial graphene/SiC substrate in order to obtain large area ofMoS2/graphene heterostructures. Using a transfer process  measurementswere used to study the electronic structure of the MoS2/epitaxial grapheneheterostructure. Due to a weak interlayer coupling the electronic structure of grapheneand MoS2 are well retained in their respective layers. However the bandalignment in the MoS2/graphene heterostructure allows under illumination acharge transfer process from MoS2 to graphene. This is revealed by adownshift of the graphene 2D Raman bands, an upshift of the A1g Raman band ofMoS2, and a strong quenching of photoluminescence (PL) ofMoS2/graphene heterostructure. Complementary we performed photocurrentmeasurements to elucidate how the presence of the semiconductor affects thephotoconductive properties of graphene. In that respect, this work may open a new way ingraphene optoelectronics by modulating the graphene photoelectric response through 2Dmaterials interfacing.One well-established method to produce high quality wafer-scale monolayer graphene is theepitaxial approach based on the graphitization of the Si face of SiC132 flakes grown bychemical vapor deposition on oxidized silicon substrates that were then transferredonto epitaxial graphene grown on SiC(0001) , we were ableto obtain several flakes with various stacking order and orientation in a singletransfer step .For the following experiments, the monolayer, bilayer and multilayer coverage wasestimated from optical analysis to be around 83%, 15% and 2% respectively.TMDCs/graphene heterostructures were made from MoSiC(0001) . ThegraiC(0001) . The CVDsolution . Afterwaubstrate . We finafer step . The MoS2/graphene/SiC heterostructure, we measured its band structure byangle-resolved photoemission spectroscopy (ARPES) at the Cassiopéebeamline of Synchrotron Soleil. The small x-ray spot size(50 × 50 μm2)allowed the measurement of the band structure of a single flake (monolayer orbilayer) forming the MoS2/graphene heterostructure. The photoelectronintensity is presented in 2 valence band. A close inspection of the K point of grapheneBrillouin zone is shown in 2 flake andgraphene underlayer is critical. In order to obtain a perfect alignment of theBrillouin zone of the two materials we need a mismatch angle of zero degree. As wecan see in 232 are not visibleindicating a non-zero mismatch angle for this flake. However we clearly see thegraphene band structure, in particular the π bands of graphene preservetheir linearity characteristic of a massless Dirac fermions signature, indicating ahigh structural quality of the MoS2/graphene heterostructure. Moreover,similar to pristine monolayer graphene, the Dirac point (ED) is locatedat 0.3 eV below the Fermi level (FL). From a linear fit, using therelation E = ħvFk, we obtain the value of the FermivelocityvF ~ 1.1 × 106 m/s,which matches the expected value for monolayer graphene on SiC. As the lineardispersion and Fermi velocity of the pristine graphene is preserved in theMoS2/graphene heterostructure, we can infer that the MoS2transfer does not affect the electronic structure of monolayer graphene in theheterostructure formation. In addition, this feature is expected theoretically sincein van der Waals heterostructures, the superposition of each layer electronicstructure constitutes a good approximation of the electronic structure of themultilayer. Differently from previous work of Diaz et al. no signature ofinterlayer hybridization2flake and the graphene underlayer. However identifying the dependence of this effecton the mismatch angle would clearly require a further work and go much beyond themain objective of this paper. 2 on epitaxial graphene,along the K-Γ-K direction in the hexagonal Brillouin zone, therespective second derivative are shown in 2 presents only one band atthe Γ point (maximum at Binding Energy (BE)~ −1.68 eV ± 0.05 eV),and this structure evolves into two branches in the case of bilayer MoS2(maximum at BE~ −1.25 eV ± 0.05 eV).This evolution is representative of the splitting of the bands due to the weak vander Waals interaction between the two MoS2 layers. The relative positionof the top of the valence bands at the Γ point in the bilayer film iscloser to the Fermi level than the one obtained from the monolayer  and Γ points. However, when considering the monolayerband structure, the top of the valence band becomes very flat near the Γpoint, leading to a direct gap at the K (K′) point. The evolution of thevalence band at the Γ point provides a straightforward method toidentify the thickness of ultrathin MoS2 films, and also proves the highquality of MoS2 transferred on epitaxial graphene. The epitaxial grapheneunderlayer does not affect the MoS2 band structure, as expected for a vander Waals heterostructure. However, we do not exclude the presence of the universalbuckled form of 2D crystal in our MoS2 layerIn order to better understand the electronic properties of theMoSonolayer . This in2/graphene heterostructure, micro photoluminescence (micro-PL) andmicro-Raman measurements are performed at room temperature (see method).To further investigate the electronic properties of mono and bilayerMoS2 todirect gap semiconductor in monolayer MoS2. The PL signal is stronglyenhanced when a direct band gap is present. The same behavior was also observed forthe as grown MoS2 on SiO2 , Γ−1 is between8–14, and presents a huge decrease as the thickness of graphene layer isreduced (~2 in the case of 5 nm FLG). In our case, we haveonly one graphene layer, meaning that as for SiO2 we can supposeΓ−1 ~ 1.Therefore, in both cases we can neglect the effect of optical interferences, anddirectly compare the PL raw data of MoS2/SiO2 andMoS2/graphene heterostructures as shown in 2/SiO2 heterostructure strain canexist due to the different thermal expansion coefficients of MoS2 and theSiO2 substrate during the MoS2 flakes growing2/graphene heterostructure38392 substrate). This phenomenonwas explored also as a function of the laser power. In 2 PL intensity evolveslinearly with increasing laser excitation412 and graphene. But in the case of graphene underlayer the PLsignal is quenched for each laser power.In the case of vertical vdW heterostructures, possible PL intensity variations couldarise from the interference effects due to different optical constants and thicknessof the different layers forming the heterostructure2 to thegraphene which hinders the recombination of electron−hole pairs createdby the photoexcitation322 and graphene , but rather to astandard hopping from an electron in MoS2 conduction band to anunoccupied state at the same energy in graphene43This phenomenon is the signature of electron transfer from MoS2/graphene heterostructure and pristine graphene layer−1. Besides thetypical second-order Raman bands that originate from the SiC substrate, the threemain structures typical of graphene are present on the pristine graphene andMoS2/graphene spectra: i) the D band (defect induced mode), ii) the Gband (in-plane vibration mode) and iii) the 2D band (two-phonon mode)2/graphene heterostructure within the wavenumberrange between 350–450 cm−1 twonew peaks are present. These two characteristic features correspond to the in-planevibration (1g) of Moand S atoms in the MoS2 film341g and 2 layers. The averageRaman spectra obtained from each layer are shown in −1,~21 cm−1 and24 cm−1 corresponding to monolayer,bilayer and multilayer (~three layers) MoS2,respectively342 on graphene and onSiO2 substrates result of a Lorentzian fitting, are narrower than those on SiO2. As in the case of the PL spectra, the upshift of the in plane Raman mode2, and confirms that the photoelectrons generated bythe Raman laser are transferred from the MoS2 to graphene. At the sametime the G peak presents an upshift. If we focus our attention on monolayerMoS2/graphene we have an upshift of~3 cm−1 ± 1cm−1.From this shift we can obtain a quantitative analysis of the level of electrondoping under illumination5357582/graphene as −1implies a change in the graphene Fermi level of ~70 meV.From these values we can calculate the variation in the n-doping of graphene usingthe relation vF = 1.1 × 106 m/s),we obtained an electron density for pristine graphene|N| ~ 7 × 1012 cm−2which increases to|N| ~ 1013 cm−2for 1ML MoS2/graphene under illumination.A detailed analysis of the Raman spectra of the monolayer and bilayerMoS2 decorated graphene) at roomtemperature while illuminating the sample with light energy(λ = 405 nm(hν ≈ 3 eV)) above theMoS2 band gap because of its thickness and limited coverage. The photocurrent generation in TMDCheterostructure generally suffers from two main limitations from an applied point ofview, which are their slow response time610.8. Such dependence isweaker than for large gain system like graphene–PbS quantum dot hybridsystem2 decorated graphenesystem over a larger range of light flux. While modulating the incident laserintensity thanks to a signal generator we can extract the frequency dependence ofthe graphene-MoS2 systemSince the spot diameter of the laser is 1 mm2 on epitaxial graphene layer. From the PL, ARPES and micro-Ramandata presented in 2/graphene heterostructurepresents a good long-range order at large scale. Our ARPES measurements on theheterostructure showed that graphene and MoS2 largely retained theiroriginal electronic structure indicating weak van der Waals interactions between thetwo crystals. The PL quenching in the MoS2-graphene heterostructure andthe upshift of the A1g Raman mode of MoS2 and the downshift ofthe 2D Raman mode of graphene confirm the charge transfer between theMoS2 and graphene layer. Our work suggests that the opticalproperties of MoS2 are strongly affected by the underlayer graphene. Thefact that an interaction is visible between MoS2 and graphene is a clearsignature of the quality of the wet transfer process and of the absence ofinterfacial contaminations. Moreover, this charge transfer can be influenced by theunderlayer graphene doping, varying the band alignment in the heterostructure, whichshould be considered in device design and fabrication. Furthermore, efficientphotoresponse was observed on large 2D MoS2/epitaxial graphene devices,opening a new way in graphene optoelectronics.In summary, we have studied the electronic properties of the wet transfer of largearea MoS2) at1550 °C to produce well-ordered atomic terraces of SiC.Subsequently, the SiC sample is heated to 1000 °C at apressure of about 10−5 mbar and thenfurther heated to 1550 °C in an Ar atmosphere. Thisgraphitization process results in the growth of an electrically active graphenelayer on top of the buffer layer, covalently bound to the substrateex-situ to perform different measurements.Monolayer graphene studied in this paper is produced via a two-step processbeginning with a starting substrate of 4H-SiC(0001)2/SiO2 samples were grown via CVD in a 1″quartz tube furnace. Microliter droplets of saturated ammonia heptamolybdatesolution were dried onto the corners of a Si/SiO2 growth substratethat had previously been coated with a layer of sodium cholate (1% solution spincoated 4000 rpm for 60 sec). Sodium cholate is a known growthpromoter, acting to increase diffusion of the molybdenum source by increasingthe surface adhesive energy relative to the adatom cohesive energy2) was used to bring sulfur vapor into the furnace fora 30 min growth period. The sample was then rapidly cooled bycracking open the furnace and sliding it downstream with respect to the quartztube.MoSThe PL measurements were carried out using a confocal commercial Renishawmicro-Raman microscope with a 100× objective and a Si detector(detection range up to ~2.2 eV). The Raman spectrameasurements were performed on the same microscope using a 532 nmlaser in an ambient environment at room temperature. To ensure thereproducibility of the data, we followed a careful alignment and optimizationprotocol. In addition, the excitation laser was focused onto the samples withspot diameter of ~1 μm and incident power of~5 mW. The integration time was optimized to obtain asatisfactory signal-to-noise ratio. We obtained Raman spatial maps by rasterscanning with 0.3 μm step size using a precision 2Dmapping stage.° and 16 meV. The meandiameter of the incident photon beam was smaller than50 μm. All ARPES experiments were done atroom temperature.The ARPES measurements were conducted at the CASSIOPEE beamline of SynchrotronSOLEIL . We used linearly polarized photons of50 eV and 90 eV and a hemispherical electron analyzerwith vertical slits to allow band mapping. The total angle and energyresolutions were 0.25For electrical measurements the samples are electrically characterized in air atroom temperature. Temporal and power dependence of the current are obtainedwhile biasing the sample and measuring the current with a Keithley 2634B assourcemeter. Illumination is ensured by a 405 nm blue laser diodewith tunable light intensity. The frequency dependence of the photocurrent ismeasured while the sample is biased using a Keithley 2634B. The output signal isamplified in a Keithley 427 current amplifier and the signal acquired on HPoscilloscope.3d5for S and Mo were used with cutoff radii (in atomic units)s = 4.3, p = 4.7,d = 5.5 (S) and s = 5.0,p = 5.6, d = 4.8 (Mo). In thisstudy we have considered standard unit cells of 3, 6 and 9 atoms to describerespectively a mono-bi- and trilayer of MoS2. Each configuration hasbeen relaxed using a sample of 32 k-points in the Brillouin zone. In case ofmultilayer MoS2, we have considered the most stable AB stacking, andthe equilibrium distance has been determined using theLCAO-S2 + vdW formalism712 are providedin First-principles calculations have been performed using a very efficient DFTlocalized orbital molecular dynamic technique (FIREBALL)676869How to cite this article: Pierucci, D. et al. Large area molybdenumdisulphide - epitaxial graphene vertical Van der Waals heterostructures. Sci.Rep.6, 26656; doi: 10.1038/srep26656 (2016).


Juvenile localised scleroderma (JLS) is a rare condition. Researchers and clinicians seeking to improve care for children and young people (CYP) with JLS face various challenges. JLS is often difficult to assess, both at disease onset and as the condition progresses. A variety of monitoring tools (mts) have been described for JLS.- To describe how mts are used and perceived by clinicians in the United Kingdom (UK).- To describe treatments used for JLS in CYP.We surveyed paediatric rheumatologists (prs) and dermatologists (dms) in the UK who manage CYP with JLS, via an an e-survey distributed to the membership of two national organisations representing these clinician groups. We asked respondents for their views and experience using 15 distinct JLS mts; about transition services and treatments used.48 clinicians  from across the UK responded. At time of survey response, dms reported managing a mean 4.2 CYP with JLS; prs 17.9. 26% dms managed all CYP in collaboration  whilst 31% prs managed all CYP in collaboration (with a DM or paediatrician). 43% of dms and 91% prs reported formal transition arrangements. 9% dms and 69% prs reported involving an adult rheumatologist in transition; 8% prs involved a DM in transition.Medical photography (100%), thermography (67%) and drawing in notes/use of pre-printed body map (both 56%) were the most regularly used mts by prs; photography (81%) drawing in notes (56%) and ultrasound (22%) were most regularly used by dms. Of 4 skin scores, the modified Rodnan skin score was the one clinicians used most frequently. However it was only used regularly by 33% prs and 3% dms. Familiarity, use  and perceived benefit of all other skin scores was low (0-40%). Laser doppler imaging, laser doppler flowmetry and 3.0T MRI were perceived useful by larger proportions (15-70%) of clinicians than reported using them  or regularly (0-11%)).Table A wide range of mts are used in the UK by prs and dms managing CYP with JLS. How these tools are accessed, used and perceived varies between and within these clinician groups. There are also differences in prescribing of treatments between prs and dms. These differences will impact on the feasibility of conducting clinical trials in JLS. Further work is needed to determine accessible and validated mts for CYP with JLS. Collaboration between dms and prs is an important factor in facilitating future high quality research and standardising care in JLS.None declared.


In this paper, we report how few layers graphene that can be produced in large quantity with low defect ratio from exfoliation of graphite by using a high intensity probe sonication in water containing liquid hand soap and PVP. It was founded that the graphene powder obtained by this simple exfoliation method after the heat treatment had an excellent exfoliation into a single or layered graphene sheets. The UV-visible spectroscopy, FESEM, TEM, X-ray powder diffraction and Raman spectroscopy was used to analyse the graphene product. The thermal diffusivity of the samples was analysed using a highly accurate thermal-wave cavity photothermal technique. The data obtained showed excellent enhancement in the thermal diffusivity of the graphene dispersion. This well-dispersed graphene was then used to fabricate an electrically conductive polymer-graphene film composite. The results demonstrated that this low cost and environmental friendly technique allowed to the production of high quality layered graphene sheets, improved the thermal and electrical properties. This may find use in the wide range of applications based on graphene. Recently, graphene as a single atomic layer of carbon has attracted much attention due to its unique nanostructure and novel thermal, electric and mechanical properties as well as its potential applications in physics, materials science, chemistry and biology . Graphen−4 m2 s −1, was about one order of magnitude lower than single layer graphene. Gupta et al [We suggest that the molecules in hand soap fluid may have similar interactions with graphite flakes to produce single or few layers graphene sheets. For testing this, hand soap—PVP (SP), as the solvent and dispersant and stabilizer, was selected to prepare a stable dispersion of the individual graphene sheets in water by using a high power sonicator. Benefits of hand soap include its low cost, established safety, non-pollutive properties and hence, a low environmental risks. In this Article, the probe ultrasonication technique produces the stable dispersions of a few layered graphene in SP solution with low defects, hence resulting in a high electrical conductivity. After wash, dry and thermal treatment, we obtained pure graphene powder, with high quality and with a low defect ratio. Kuang et al and other research groups –31 have ta et al  found thta et al –36.To outa et al –39. The The pure graphite powders  and Polyvinyl pyrrolidone  were purchased from Sigma- Aldrich company.Solvent: Liquid hand soap  was dissolved in deionized (DI) water to create a solution of hand soap-DI water, 0.5 wt%. For exfoliation and dispersion of graphite in the solution, a probe ultrasonic homogenizer (UH)  was used with high intensity. The sonication probe tip was provided in continuous mode for 60% amplitude modulation.The mixture, SP, containing 1g PVP with 400 ml of hand soap-DI water (0.5 wt%) was stirred for 1 h, then, 1 g of pure graphite (GT) powder was added to the SP solution to form graphite solution. The resulting solution was tip sonicated for 3 hrs, and then was left to stand for 24 hrs to separate the large graphite aggregates to the bottom. The homogenous dark dispersion remained suspended for at least five months after preparation without any significant precipitation  was dropped on the substrate during the spin coating process at 1000 rpm for 30 s, and finally, the film was subjected to heat treatment at 100°C under vacuum for 48 hrs to remove the solvent remnants and produce the multi-layered graphene. The quartz substrates were washed by distilled water and acetone and then placed in an ultrasonic cleaner for 30 min for the cleaning process.The optical absorption of the diluted solution before and after ultrasonic treatment was analysed using an UV−vis spectrophotometer (Perkin Elmer model Lambda 900). The powder morphology of the pure GT, the resulting graphitic powder and films were determined by field-emission scanning electron microscopy  and transmission electron microscopy  and X-ray diffraction (XRD)  equipped with a Cu Kα  radiation source. Raman spectroscopy was performed  which was used to analyze edge defects and estimate layer thickness. X-ray photoelectron spectroscopy (XPS) measurement was carried out using a Multifunctional XPS (Shimadzu Axis Ultra DLD). The electrical resistance of graphene composite film was characterized by four point probe measurement using a Keithley 2401 source meter.V was determined by the thickness and thermal diffusivity of sample, V(f) = V0eσL− where V (f) is the complex PE signal, σ is the complex TW diffusion coefficient and L is the liquid sample thickness. From a relation σ = (1 + i)/μ, the thermal diffusion length of sample is μ = (α/π f)1/2, where f is the modulation frequency, and α is the thermal diffusivity of sample. The amplitude and phase of the signal are respectively, [The experimental TWC technique for measuring the liquid thermal diffusivity was utilised as reported previously . In thisctively, |V(f)|=VS = L(π/α)1/2.From the graph of the ln(amplitude) and phase as a function of the square root of the frequency, the thermal diffusivity of liquid sample can be calculated from the slopes of The extent of exfoliation for graphite could be increased with increased ultrasonication time, due to the high energy activation of ultrasonication . This acFor investigating the graphene exfoliated by sonication treatment, the optical absorption of the SP-G solution before and after sonication of 3 hrs was analysed, TEM was employed to show the quality and morphology of the SP-G solution prepared with the ultrasonic method. The XRD patterns of pure GT, and the prepared samples without (SP-G) and with (G) heat treatment are displayed in -1, which corresponded to the D band and G band, respectively. The D band at 1350 cm-1 corresponds the breathing mode of sp2 carbon atoms, and it is absent without the presence of defects and edges in the graphitic structure [ID/IG = 0.15), while as expected a small ratio (GID/IG = 0.35) is found for the exfoliated graphene sheets. We propose that the ratio of the defects GID/IG to the defect GTID/IG (i.e. M = GID/IG /GTID/IG) can be used to quantitatively estimate the graphene sheets. We suggest using the ratio of the intensity of the D: G band ratio of graphene sheets, divided by the intensity of the D: G band ratio of the graphite source. This relationship  allows the measurement of the mean defects of the graphene sheets from the graphite used to produce the graphene. The edge defects factor of the exfoliated graphene in -1) is an effective means to determine the number of layers in the graphene sheets. As seen in Raman Spectroscopy is a powerful tool to determine the number of layers present along with the quality of the graphene present (relative defects in graphene) with a faster throughput than other techniques such as AFM . Fig 7A tructure . The pur2 (C-C) and consisted of peaks corresponding to carbon bound (C–C) at ~284.5 eV and (C-O) at around 286 eV, respectively [To further evaluate the quality and also the oxygen containing groups in the sheet, XPS analysis was performed for the pure GT and the G powder obtained by sonication after heat treatment. The high resolution of the sharp and large carbon 1s (C1s) peak of GT and G represented the graphitic carbon spectively ,51  Where αn, ρn and Cn are the thermal diffusivity, density and specific heat capacity of the nanofluids. It can be seen that in low concentration of grapheme obvious enhancement in thermal diffusivity can be expected with similar enhancements in thermal conductivity. To the best of our knowledge, there are no reports on thermal diffusivity of the exfoliated graphene nanofluids. Therefore, this method to measure thermal diffusivity of the graphene nanofluids may provide a new Insight to measure different properties of graphene nanofluids. This enhancement is in agreement with other reports in the literature for thermal conductivity of the graphene composites . While, These results confirm that the solution containing graphene had a large scale exfoliation into single or few layer of graphene that can be good candidates for high thermal conduction in graphene or graphene composite devices. The physical and chemical properties of graphene depend on its crystalline structure along with the number of layers. The combination of TEM observations along with, UV-vis absorption, conductivity measurements, and thermal properties clearly indicate a dispersed graphene solution with high stability, using this environmental solvent. The lack of any individual single layer graphene sheets suggested that the graphite cannot be fully exfoliated. Separation of graphene layers depends on the mechanical disruption of the graphene stacking by ultrasonication ,18. In oIn conclusion, a simple liquid exfoliation method of producing layered graphene with a low defect ratio from pure graphite powder was obtained by a sonication treatment in a mixture of SP surfactant in DI water. The presence of SP solution in this technique allowed an excellent dispersion of graphene sheets with a high stability, even after 6 months. In the thermal diffusivity of the SP solution with exfoliated graphene demonstrated a precise enhancement (by a factor of 3). This enhancement compared favourably to other graphene-loaded thermal conductive graphene composites. The fabricated film from SP-G solution containing graphene was utilized to make an electrical conductive polymer-graphene film composite, the sheet resistance was found to be 3 order magnitude lower than that of SP solution. Graphene with a precise dispersion was obtained from this technique, without any oxidization or significant defect formations. The grapheme produced is very promising for the development of the graphene based biomedical and electrical applications.S1 FigMore information about TWC technique(DOCX)Click here for additional data file.S2 FigThe frequency behavior of the Ln amplitude and phase of PE signal obtained from DI water (a) ln (amplitude), and (b) phase of PE signal versus the square root of the frequency for one of DI water.(DOCX)Click here for additional data file.


Due to the interactions from the substrate, a much large exchange field (about 280 meV) and an enhanced Rashba spin-orbit coupling are induced in graphene, leading to a topologically nontrivial QAH gap opened in the system. The avenues of enhancing the nontrivial gap are also proposed, from which nearly a gap one order large is achieved. Our work demonstrates that this graphene-based heterostructure is an appropriate candidate to be employed to experimentally observe the QAH effect and explore the promising applications.Quantum anomalous Hall (QAH) effect, with potential applications in low-power-consumption electronics, is predicted in the heterostructure of graphene on the (001) surface of a real antiferromagnetic insulator RbMnCl Through the proximity effect, the topmost FM Mn plane in RbMnCl3 may induce a strong FM exchange field in graphene, a key factor to achieve the QAH effect. This material can be grown easily by molecular beam epitaxy methods in experiments, whose (001) surface also has a hexagonal structure293 could be a suitable substrate for graphene to gain the QAH state. Based on density-functional theory (DFT) and Wannier function methods, we demonstrate the QAH effect in graphene and (001) RbMnCl3 heterostructure. A much larger exchange field (about 280 meV) is obtained in graphene. Berry curvature, Chern number, and anomalous Hall conductivity are calculated to confirm the topologically nontrivial band gap opened by the SOC. Our results demonstrate that this heterostructure is an appropriate candidate system for exploring the QAH effect and corresponding electronic applications.In this work, we tend to achieve the QAH state in the heterostructure of graphene on the (001) surface of a real antiferromagnetic (AFM) insulator RbMnCl3 compound has a hexagonal structure with the experimental lattice constant of a = 7.16 Å3 with Mn terminal, as illustrated in 3 lattices is about 2.5%. In our calculations, the graphene lattice is shrunk to fit the substrate lattice. The C atoms in graphene and the topmost Mn, Rb, and Cl atoms in the substrate were all relaxed to obtain the stable structure of the heterostructure. The magnetization of the Mn atoms close to the graphene in the heterostructure is found to be along the c-axis. Its energy is lower than that of the magnetization in the ab plane by 0.25 meV per unit cell. Three possible adsorption configurations were considered: the hollow, bridge, and top sites, which stand for the topmost Mn atoms of the substrate right below the hexagon centers (hollow sites), the hexagon edges (bridge sites), and the hexagon corners (top sites) of the graphene, respectively. The relative stability of the three configurations is judged by comparing their adsorption energies, defined as Eb = EG + ER − EG+R, where EG, ER, and EG+R represent the total energies of the bare graphene supercell, the bare RbMnCl3 film, and the graphene/RbMnCl3 heterostructure, respectively. The hollow site is found to be the most stable configuration, displayed in 0 (d0 = 2.03 Å) between the graphene plane and the topmost layer of Mn atoms in the substrate after the geometry optimization.The RbMnCl3 substrate, it is magnetized by the proximity coupled effect with the topmost ferromagnetic Mn layer. 3 film. We find that the Dirac bands of graphene are just located inside the bulk band gap of the RbMnCl3 film. Obvious spin splitting appears in the bands around the Dirac point. Since the K and K’ Dirac points of the pristine graphene are both folded into the Γ point of the 3 × 3 supercell of graphene, each spin-up and spin-down Dirac bands around the Γ point are twofold degeneracy. Magnifying the spin-polarized π bands around the Dirac point at the Γ point (12−3  meV)d states in the substrated orbit to the Dirac bands increases, leading to a larger exchange field and Rashba SOC. Thus, the opened bulk band gap could increase slightly to about 2.0 meV in this case. Γ point , we can The mechanism of the gap opening can be understood by a tight-binding (TB) model of graphene with a TM adsorbed at the hollow site in a 3 × 3 supercell. We consider the TM-induced on-site potential, magnetic exchange field, and Rashba SOC on the six carbon atoms nearest to the TM adatoms. The TB Hamiltonian can be written as719i, z axis, m(n) refers to the sites nearest to the TM atom, andm to site n. The first term represents the nearest-neighboring hopping with amplitude t = 2.6 eV, the second term is the intrinsic spin-orbit coupling. The last three terms correspond to the TM-induced extrinsic Rashba SOC, on-site potential and magnetic exchange field, respectively. For the TM-induced intrinsic SOC is usually much smaller than the induced Rashba SOC, we neglect the intrinsic SOC term by setting where ulations .k) is calculated by3132333435The Berry curvature and Chern number are calculated to identity whether the SOC-induced gap is topologically nontrivial. The Berry curvature Ω. For two-dimensional systems, the calculated Berry curvature is the z-component value. The Chern number C and Hall conductivity σxy can be obtained by integrating the Ω(k) over the first BZ, as 3 heterostructures. The peaks of the Berry curvature are primarily located at the k points with the SOC-induced band gaps. The corresponding distribution of Ω(k) in the two-dimensional (2D) momentum space is given in k) distributes very close to the Γ point in the first BZ. Integrating the obtained Ω(k) over the first BZ, the Chern number of C = 2 is obtained. This integer Chern number (C = 2) characterizes a quantized Hall conductivity and confirms the topological non-triviality of the gap opened by the SOC interaction. Two chiral edge channels will arise on each side of the sample. Therefore, the QAH effect can appear in the graphene/RbMnCl3 heterostructure. Since the nontrivial gap is opened below the Fermi energy (EF), hole doping is needed to observe this exotic effect in experiments. Our first-principles calculations indicate that doping one hole per unit cell could move the EF to the crossing points of spin-up and spin-down bands, as shown in 14 cm−2, which can be experimentally achieved through currently advanced gating technologies. For instance, employing an electrolytic gate, the doping concentration in graphene can be adjusted up to 4 × 1014 cm−2 in experimentsF.where the summation is over all the 3 substrate is strengthened, the exchange field and Rashba SOC in the graphene can be enhanced. To increase the interaction, we reduce the distance between the graphene sheet and the substrate, by applying an externally vertical stress to the heterostructure. As shown in g can be almost doubled (2.1 meV). In experiments, the exerted stress, however, cannot be too large. The structure of the systems may be destroyed under a very large strain.If the interaction between the graphene and RbMnCl3 heterostructure with Hf atoms adsorbed at the hollow site of the graphene. A relatively large band gap of about 9.0 meV appears in the system. Thus, compared to the original gap, nearly one order large gap is achieved by adding Hf atoms into the system. The origin of this large band gap comes from the enhanced Rashba SOC due to the adsorption of Hf atoms, as reported in Refs. k) and its corresponding 2D distribution in momentum space are illustrated in F is adjusted in the gap. The Ω(k) peaks are also localized around the SOC-induced band gap near the Γ point. The intrinsic anomalous Hall conductivity σxy as a function of the EF is plotted in F passes the SOC-induced bulk band gap, confirming the existence of the QAH effect in this proposed system. The Hall conductivity in F is larger than −1.24 eV, ascribing to the spin-direction reversal of the main bands around this energy point  surface of an AFM insulator RbMnCl3 film, opposite to the graphene interface, was passivated by hydrogen atoms. A vacuum space of larger than 15 Å was used to avoid the interaction between two adjacent heterostructure slabs. For the structural relaxation, the C atoms in graphene and the topmost Mn, Rb, and Cl atoms in the substrate were allowed to relax until the Hellmann-Feynman force on each atom was smaller than 0.01 eV/Å. All calculations were carried out with a plane-wave cutoff energy of 550 eV and 12 × 12 × 1 Monkhorst-Pack grides were adopted for the first Brillouin zone (BZ) integral. To take into account the correlation effects of Mn 3d electrons, the GGA + U methodThe geometry optimization and electronic structure calculations were performed by using the first-principles method based on density-functional theory (DFT) with the projector-augmented-wave (PAW) formalismHow to cite this article: Zhang, J. et al. Quantum Anomalous Hall Effect in Graphene-based Heterostructure. Sci. Rep.5, 10629; doi: 10.1038/srep10629 (2015).