Functionalized graphene sheets were customised as a matrix for covalent immobilization of Fenugreek β-amylase using glutaraldehyde as a cross-linker. The factors affecting the process were optimized using Response Surface Methodology based Box-Behnken design of experiment which resulted in 84% immobilization efficiency. Scanning and Transmission Electron Microscopy  and Fourier Tansform Infrared (FTIR) spectroscopy were employed for the purpose of characterization of attachment of enzyme on the graphene. The enzyme kinetic studies were carried out for obtaining best catalytic performance and enhanced reusability. Optimum temperature remained unchanged, whereas optimum pH showed shift towards acidic range for immobilized enzyme. Increase in thermal stability of immobilized enzyme and non-toxic nature of functionalized graphene can be exploited for production of maltose in food and pharmaceutical industries. It is essential for the generation of maltose from cereal grain starch, thus making its role important in mashing and brewing process. The saccharifying activity is also exploited in bread making and in the use of malt as an additive in food stuffs. Maltose has properties such as mild sweetness, good thermal stability, low viscosity in solution and lack of colour formation, which accounts for its applicability in pharmaceutical dispensing and in the production of non-digestible sweetner . Major use of β-amylase is in being the exclusive source of carbon in the production of diphtheria pertussis tetanus vaccine. The characteristic property of enzyme of yielding a single product maltose, is utilized in the structural studies of starch and glycogen Enzymes being highly selective catalyze efficient reactions with few by products, thus they are environmentally favourable alternative to conventional chemical synthesis, particularly in the food and pharmaceutical industries where high reaction selectivity is essential. But, lack of long term operational stability, difficult recovery and reuse of the enzyme, limit their industrial applications. These limitations can often be overcome by immobilizing them on suitable matrices. The interaction between the enzyme and carrier material governs the properties of immobilized enzyme, thus, influencing the chemical, biochemical, mechanical and kinetic properties. The property of immobilized enzyme is also affected by particle mobility which is governed by particle size and solution viscosity Nano-materials and nano structures generally provide large surface area, low mass transfer resistance and high enzyme loading capability, which enables better interaction with the enzyme, resulting in increase in immobilization efficiency and enhancement in long term storage and recycling ability of the enzyme. Nanomaterials can also be engineered to present multiple surface functional groups for interacting with biomolecules. A wide range of nano materials have been studied and their positive effect on enzyme was shown through immobilization process Various graphene based nano materials have been used to fabricate functionalized biosystems integrated with nucleic acids, peptides, proteins and even cells. Graphene is a free standing 2D crystal with one atom thickness. This allotrope of carbon comprises layers of six atom rings in a honey combed network and can be conceptually viewed as a true planar atomic macromolecule. Graphene also gets benefitted by biological modification which improves its biocompatibility, solubility and selectivity. Proteins can exfoliate and modify graphene due to presence of various functional groups, through physical adsorption or chemical bonding Response surface methodology (RSM) is a widely practiced approach for the production and optimization of different industrially important biotechnological and biochemical products. It is a collection of statistical and mathematical techniques useful for developing and optimizing processes. RSM is a method for constructing global approximations to system behaviour based on results calculated at various points in the design space. It also determines effect on specific response resulting from the changes in the level of the factors.The choice of experimental design is affected by the shape of the experimental region, which is determined by the ranges of the variables. The Box-Behnken design is an independent rotatable or nearly rotatable quadratic design. It excludes the corners, where all variables are simultaneously at the maximum levels. The use of the Box-Behnken design is popular because it is an economical design and requires only three levels for each factor. Therefore, it requires few experimental combinations of the variables β-amylase on functionalized graphene using RSM based Box-Behnken design of experiment and to study the effect of immobilization on kinetic parameters. Immobilized enzyme was characterized using SEM and TEM. The chemical interactions and bonding involved during immobilization were characterized by FTIR spectral analysis.The basic objective of the present study include immobilization of Fenugreek Trigonella foenum-graecum) were purchased from local market. The chemicals for buffers preparation were of Analytical or Electrophoresis grade from Merck Eurolab GmbH Damstadt, Germany. All other chemicals and reagents were purchased from Sigma Chem. Co. Milli Q  water with a resistance of higher than 18 MΩ cm was used all throughout the experiment.Dry seeds of fenugreek (β-Amylase was purified from dry seeds of fenugreek (Trigonella foenum-graecum) and had a specific activity of 732.59 units/mg. Homogeneity of the purified preparation was checked by SDS-PAGE Protein concentration was determined by using Bradford's method (1976) Enzyme activity was measured by following Bernfeld's method (1955) β-amylase were incubated with 0.5 mL of starch solution (1%) prepared in 50 mM sodium acetate buffer, pH 4.5 for 3 min at 37°C, followed by centrifugation at 5000 rpm for 2 min at 4°C. Supernatant was pipetted out in test tube and 1 mL of 3,5 dinitrosalicylic acid was added, followed by placing it in boiling water bath for 5 min. It was diluted with 10 mL of Milli Q water before recording absorbance at 540 nm.Graphene sheets cross-linked with Fenugreek β-amylase is defined as the amount required for release of 1 µM of β-maltose per unit at 30°C and pH 5.0, under the specified condition.One unit of The immobilization efficiency was calculated using following formula:Enzyme units were calculated using starch as substrate, under standard assay conditions as described earlier. The immobilized protein was determined by subtracting the protein estimated in supernatant after immobilization from the total amount of protein used for immobilization.2SO4, HNO3 and KClO3 at room temperature under constant stirring condition. The graphite oxide solution obtained was washed with distilled water and 10% HCl solution for removal of sulphate and other ionic impurities. It was then dried under vaccum at 80°C. The graphite oxide powder was placed in an alumina boat and inserted into a 1.5 m long quartz tube with outer diameter of 25 mm. The sample was flushed with Ar gas for 15 min followed by insertion of quartz tube into a tube furnace pre heated at 1050°C and was kept in it for 30 sec. The functionalized graphene sample was allowed to cool down to room temperature under Ar gas flow. Thermally exfoliated functionalized graphene thus prepared, was light weighted shiny black powder which differed in appearance from brownish graphite oxide. The sample prepared was suspended in double distilled water and was sonicated at room temperature for 10 min. It was then left undisturbed for 30 min for the settlement of the larger non exfoliated flakes and the suspended functionalized graphene was collected.Functionalized graphene sheets were prepared by thermal exfoliation of grahite oxide, following the protocol described by Staudenmaier The enzyme was coupled to the functionalized graphene with the help of glutaraldehyde as a cross linker. A preparation of 1 mg/mL of graphene was made by dissolving functionalized graphene in 50 mM sodium phosphate buffer (pH 7.0), which was divided into 17 aliquots according to the design of the experiment . These a−1 wave number. A good signal to noise ratio was achieved with 100 scans of each sample.The characterization studies of functionalized graphene sheets (both native and coupled) wereperformed by Transmission Electron Microscope , Scanning Electron Microscope  and Fourier Transform Infrared Spectroscopy . TEM studies were done by placing a drop of sample on electron microscope 200 mesh copper grid and the water was allowed to get evaporated for complete dryness of the sample, followed by its loading into machine. SEM studies were carried out by sprinkling the samples on the stub having layer of silver glue for striking the particles. Secondary electron imaging mode was used for obtaining fine structural details. FTIR was performed in the range of 650 to 4000 cmβ-amylase onto functionalized graphene were: amount of graphene ; concentration of glutaraldehyde  and amount of enzyme . The quadratic polynomial equation relating the variables to the responses was used for calculation of the mathematical relationship:The experiment was designed using Box-Behnken design for analysis and optimization of the factors  which influence the process. The initial values of the factors affecting the process of immobilization were determined by carrying out some preliminary experiments (data not shown). ‘Design Expert’ software  was used for experimental design and analysis. The Box-Behnken design used for fitting the quadratic model led to the experiment consisting of 17 trials and all the factors studied were varied at three different levels. The variables and their levels selected for carrying out immobilization of Fenugreek iY is the predicted response, iXjX are input variables which influence the response variable Y; oβ is the offset term; iβ is the ith linear coefficient; iiβ the ith quadratic coefficient and ijβ is the ijth interaction coefficient. The co-efficients were estimated by performing the 17 trials and the generated mathematical model was validated by carrying out experiment at given optimal condition. The model was statistically analysed for evaluation of the anlaysis of variance (ANOVA), which determines the validity of the model based on Fischer's F-test, associated probability, correlation coefficient R and lack of fit test.Where, In all cases, control experiments using soluble enzyme were carried out.β-amylase was studied in the pH range of 3.0–9.0 . 50 mM buffers were used in each case and assay procedure was performed under standard conditions. The optimum temperature was studied by carrying out assay procedures at temperatures between 25–75°C. The thermal stability was determined by incubating the enzyme at 55°C for different time intervals, followed by residual activity assay under standard condition. Starch concentration was varied in the range of 0.5–10 mg/mL for observing effect of substrate concentration on immobilized enzyme. Data thus obtained was used for calculation of mK and maxV from Lineweaver-Burk plot. For the assessment of reusability, the immobilized enzyme was used for carrying out assay procedure repeatedly. After each assay, the immobilized enzyme was washed with 50 mM sodium phosphate buffer, pH 7.0. The aliquots of functionalized graphene coupled with enzyme were stored at 4°C and the residual activity was determined during 4 months storage.The effect of pH on immobilized Fenugreek +0.5).For obtaining optimal conditions for enzyme immobilization, RSM was employed. Central Composite design and Box-Behnken design are the two designs of experiment commonly utilised in RSM. A comparative study between the two designs of experiment was done by Ferreira et al (2007) µg) and C, enzyme (µg).These experimental results formed the base for building of model for obtaining maximum immobilization response as presented in For establishing the adequacy and significance of predicted quadratic model, ANOVA was performed . The ModSystematical characterization of the enzyme immobilized on functionalized graphene was done by carrying out SEM and TEM, followed by FTIR spectroscopy. Bright field TEM images showed typical transparent sheets of functionalized graphene at 50 nm resolution, the characteristic Selected Area electron Diffraction (SAD) pattern obtained is shown in inset of −1 (C = C band), which has been assigned to skeletal vibrations of graphitic domains. Functional groups present in graphite oxide  are utilized in its further functionalization. The peak at 1740 cm−1 is characteristic of −COOH (−C = O in carboxylic acid) which is known to appear after execution of oxidation process. The O-H stretching mode of −COOH groups and −OH groups attached on the surface of functionalized graphene leads to appearance of peaks of O-H at ∼1400 cm−1 together with the band at 3450 cm−1. The vibration of C-O in carboxylic acid groups or epoxy groups results in appearance of the peak at ∼1054 cm−1. The cross linking of functionalized graphene has also been characterized by FTIR. Peaks corresponding to the C-H stretch modes of the cross linked molecule was located at 2927 and 2858 cm−1. An increase in the relative intensity between peaks at 1054 cm−1 and 1630 cm−1 signifies the increase in the intensity of C-O stretch mode and suggests that cross linking between functionalized graphene and glutaraldehyde has taken place. The hydroxyl groups can react with aldehyde through formation of hemiacetal structure. The aldehyde functional groups present in the molecule of glutaraldehyde, react with the sheets of functionalized graphene as a cross-linker. Glutaraldehyde binds to the −OH group of the graphite oxide through its one arm containing −CHO group and attaches to the enzyme via lysine amino group through its another −CHO group. Glutaraldehyde is also known to play important role in the self-assembly of graphite oxide nanosheets in aqueous solution, as the self-assembly of nano sheets occur in more ordered fashion in presence of little amount of glutaraldehyde −1, representing carbonyl amide I bond whereas bands at 1481 and 1567 cm−1 were observed due to amide II bonds suggesting immobilization of enzyme on functionalized graphene.The natural graphite shows a characteristic peak located at 1625 cmAn alteration in kinetic parameters is often brought about by immobilization process due to changes in the microenvironment of immobilized enzyme β-amylase with respect to activity and stability. The optimum pH of the immobilized Fenugreek β-amylase was found to be 4.5, while the soluble enzyme had pH optima at 5.0. The extent of displacement of optimum pH is influenced by interaction between the enzyme and the matrix and also by the properties of matrix. Hydrolytic activity was observed in a wider pH range by the immobilized enzyme. Compared to soluble enzyme, immobilized one showed better activity in the alkaline range (pH 7.0–9.0). β-Amylase from sweet potato immobilized on chitosan and α-cyclodextrin also showed similar results −1 and 0.0213 min−1, respectively. The immobilization support generally provides protecting effect against enzyme deactivation occurring at high temperature. The immobilization process affects the conformational flexibility and results in increase in enzyme rigidity, which is observed in form of stability towards denaturation resulting from rise in temperature β-amylase from soybean showed a small increase in thermostability on being immobilized on chitosan beads β-amylase immobilized on functionalized graphene showed better activity at higher temperatures as compared to β-amylase immobilized on phenyl boronate agarose mK value showed slight change from 1.56 to 1.82 mg/mL on being immobilized on functionalized graphene. These differences can be related to the change in microenvironment resulting from the enzymatic hydrolysis of substrate.Industrial applicability of any enzyme is judged by the knowledge of its storage stability. Stabilization of enzyme by immobilizing it on suitable matrix has been the subject of considerable research, as the intra and inter molecular cross-linking leads to a more rigid molecule that can resist conformational changes. The immobilized enzyme was found to be quite stable when stored at 4°C for 120 days with retention of 60% residual activity, whereas soluble enzyme showed residual activity of 20% during storage for same duration (β-amylase immobilized on functionalized grapheme nanosheets (Another important parameter for industrial application is reusability. Reusability in terms of retention of more than 76% enzymatic activity after 10 repeated uses was shown by Fenugreek nosheets . Repeateβ-Amylase covalently attached onto the functionalized graphene showed wider range of pH operability, better thermal stability and greater storage ability as compared to soluble enzyme, making it suitable for production of maltose at industrial scale.Optimization of immobilization using RSM based Box-Behnken design of experiment was proved to be efficient and economical, due to requirement of less number of experiments to be performed. Fenugreek


A predictive model is proposed that quantitatively describes the synergistic behavior of the electrical conductivities of CNTs and graphene in CNT:graphene hybrids. The number of CNT-to-CNT, graphene-to-graphene, and graphene-to-CNT contacts is calculated assuming a random distribution of CNTs and graphene particles in the hybrids and using an orientation density function. Calculations reveal that the total number of contacts reaches a maximum at a specific composition and depends on the particle sizes of the graphene and CNTs. The hybrids, prepared using inkjet printing, are distinguished by higher electrical conductivities than that of 100% CNT or graphene at certain composition ratios. These experimental results provide strong evidence that this approach involving constituent element contacts is suitable for investigating the properties of particulate hybrid materials. Carbon nanotubes (CNTs) and graphene have been intensively studied for various applications123467912151619et al. suggested that the synergism of the conductivities is derived from the graphene particles filling the space between the CNTs and thereby providing a lower junction resistance between the graphene and the CNTs. These two explanations for the synergistically improved electrical conductivity of CNT:graphene hybrids, i.e., CNTs acting as conducting bridges between the graphene flakes and graphene particles filling the space between CNTs, have the common feature of an increased number of contacts between the graphene particles and the CNTs. This study aimed to establish theoretical and experimental explanations for the synergistic effect. CNT:graphene hybrids prepared by simple dispersion and a film processing method were used in the experimental work.CNT:graphene hybrids have been studied in many researches due to its enhanced properties than that of graphene and CNT itself. In some work, it was found that the thermal conductivity increases by hybridizing CNT/graphene1222232425262728302730323323242528Various processing methods have been developed to make CNT:graphene hybrid films. Vacuum filtration was the first solution approach24122023151738This paper presents a predictive model for quantitatively explaining the synergistic behavior of CNT:graphene hybrids. A quantitative analysis carried out using the model demonstrated that the hybrids can exhibit higher electrical conductivities than pure CNT or graphene films at certain composition ratios. CNT:graphene hybrids prepared by inkjet printing were used to validate the predictive model.A predictive model was developed to explain the synergistic behavior of graphene and CNT electrical conductivities in CNT:graphene hybrids. The conductivity of these hybrids is governed by inter-particle contacts. A random stick network model can be used to estimate the electrical conductivity of a CNT filmns is the number of conductive sticks and nj is the number of their junctions, i.e., the number of stick-to-stick contacts. Here, b is a constant, Gs is the stick conductance, and Gj is the junction conductance. Because the conductivities of individual CNTs and graphene are much higher than the conductivity of their junctions (contacts)42where et al. defined the orientation of fibers in space using two angles , with the assumed geometries and sizes of graphene particles and CNTs, then the total number of CNTs and graphene particles in a specific volume is k. Equation Equation α and β are positive constants. In this system, 56.7 wt% is the CNT fraction that can bring about the maximum number of contacts in the modeled CNT:graphene hybrids. At this concentration, the proportion of graphene-to-CNT contact is the largest (75.0%), followed by CNT-to-CNT (19.2%), and graphene-to-graphene (5.8%) contacts. A synergistic behavior of CNT:graphene hybrids can be observed at this CNT content.where Next, the effect of graphene particle size on the number of contacts was investigated. For simplification, the size of the graphene particles was assumed to be where x μm, the number of CNTs can be expressed as The effect of CNT size on the number of contacts can be investigated by changing either its diameter or length. For simplicity, the mass of a CNT is assumed to be independent of its diameter and proportional to its length. By changing the length from 5 to 5The number of contacts that graphene and CNT particles can make in their hybrids can thus be calculated using a simplified geometry for the CNT and graphene. The model demonstrates that the condition of maximum contact occurs at a specific concentration of both materials. We can deduce that properties sensitive to such contact, e.g., electrical conductivity see Eq. , will beoC)There are several requirements for inks to be printed acceptably by an inkjet printer. The inks should have relatively low viscosity  and low volatility . The graphene particles were randomly deposited on the paper . The PAN–1, respectively. Although pure graphene has a higher electrical conductivity than pure CNTs4239The electrical conductivity of printed ink was measured using the four-point probe method. The electrical properties of the CNT:graphene hybrid inks are shown in The number of CNT-to-CNT, graphene-to-graphene, and graphene-to-CNT interparticle contacts was calculated using an orientation density function of graphene and CNT in a hybrid assembly. CNTs and graphene particles were assumed to be straight cylinders and disks, respectively. The probability of each type of contact was then calculated, from which the number of contacts was obtained. The total number of contacts followed a parabolic equation as a function of CNT concentration, suggesting the existence of a maximum at a specific composition ratio. This indicates a synergistic effect of the properties of the CNT:graphene hybrids, in particular those properties that are contact-dependent. This synergistic effect was experimentally investigated by using inkjet printing to prepare CNT:graphene hybrids. Hybrid inks, prepared at a certain composition ratio, had higher electrical conductivities than those of pure CNT or graphene, i.e., a synergistic effect. These findings support the use of our statistical approach to investigate the properties of particulate hybrid materials involving contacts of the constituent elements.Our model was aimed to estimate the electrical conductivity of CNT:graphene hybrids. However, we have found out that the percolation threshold of CNT composites can be estimated using the equations derived in our work. In previous research, the critical volume fraction ensuring the percolation was estimated by calculating the probability of one cylinder intersecting at least two other cylinders. In a work by Munson-McGee, the critical volume fraction was chosen when the probability that one cylinder intersecting at least two other cylinder is 0.5The size distribution of graphene particles and CNTs can be considered in the calculation of the number of contacts through size distribution function. The number of CNT-to-CNT, graphene-to-graphene, and graphene-to-CNT contacts can be derived as follows  was then added to each PAN solution. The graphene solutions were stirred and then ultrasonicated. Then, the solutions were centrifuged at 5000 rpm for 15 min and the supernatant was vacuum-filtered through filter paper having a pore size of 1.2 μm . CNT and graphene inks were prepared in the same manner. The separate graphene and CNT inks were then mixed together in various proportions to make different CNT:graphene hybrid inks.Graphene particles , multi-walled carbon nanotubes , poly(acrylonitrile) , and N,N-dimethylformamide  were used to prepare CNT and graphene hybrid inks using the following process. First, PAN solutions were prepared at 0.25, 0.5, 1, and 2 wt% concentrations in DMF (70 g). The solutions were stirred on a hot plate at 90 °−1 at 25 °C to adjust the printing parameters. The average size of the particles in an ink was measured using a dynamic light scattering spectrophotometer . The zeta potential was also measured using an electrophoretic light scattering spectrophotometer  to evaluate the degree of dispersion of the particles. TGA was carried out to calculate the amount of graphene, CNT, and PAN in the inks.The viscosity of each solution was measured using a rheometer  at shear rates of 4.642, 10, 21.55, 46.42, 100, 215.5, and 464.2 s2 with a drop spacing of 25 μm, varying the number of printings from 1–25. The thicknesses and morphologies of the printed inks were investigated using AFM and SEM, respectively. The electrical properties of the printed inks were measured using the four-point probe method.An inkjet printer  was used to print the prepared inks on a substrate. The droplet formation of the inks was controlled by changing the cartridge settings, including the jetting voltage, waveform, meniscus control, and temperature. Photo paper was selected as the substrate. The inks were printed in a square pattern 7 × 7 mmHow to cite this article: Shim, W. et al. Optimally conductive networks in randomly dispersed CNT:graphene hybrids. Sci. Rep.5, 16568; doi: 10.1038/srep16568 (2015).


Angiogenesis is an important process involved in the pathogenesis of diffuse parenchymal lung diseases. The aim of the study was to compare the angiogenic profile of patients with sarcoidosis and idiopathic pulmonary fibrosis (IPF) based on analysis of circulating factors.Serum concentrations of angiopoietin-2 (Ang-2), follistatin, granulocyte-macrophage-colony stimulating factor (GM-CSF), interleukin-8 (IL-8), platelet derived growth factor-BB (PDGF-BB), platelet endothelial cellular adhesion molecule-1 (PECAM-1) and vascular endothelial growth factors (VEGF) were measured in the patients and the healthy subjects.Serum concentrations of G-CSF, follistatin, PECAM-1 and IL-8 were significantly higher in the IPF patients in comparison with the control group and the sarcoid patients. PDGF-BB concentrations were also significantly higher in serum of IPF patients than in sarcoid patients, but not than in the controls. In contrast, Ang-2 and VEGF concentrations did not differ significantly between the three groups. In the sarcoid patients, irrespective of the disease activity or the radiological stage, serum concentrations of these cytokines were similar to the control group.These results indicate that differences may exist in angiogenic activity between patients with parenchymal lung diseases. In contrast to sarcoidosis, IPF is characterized by a higher serum concentration of different molecules involved in the angiogenic processes . The interstitial lung diseases are a heterogeneous group of diffuse parenchymal lung diseases, including sarcoidosis and idiopathic pulmonary fibrosis (IPF). IPF is characterized by tissue damage and exuberant repair with an aberrant wound-healing response leading to a severe disruption of the pulmonary architecture . On the Angiogenesis, defined as the process of growth of new blood vessels, plays a pivotal role in wound healing and may contribute to the fibroproliferation and extracellular matrix deposition observed in IPF and in advanced stages of sarcoidosis –5.Among various markers of angiogenesis, angiopoietin-2 (Ang-2), follistatin, granulocyte-macrophage-colony stimulating factor (GM-CSF), interleukin 8 (IL-8), platelet derived growth factor-BB (PDGF-BB), platelet endothelial cellular adhesion molecule-1 (PECAM-1) and vascular endothelial growth factor (VEGF), seem to be involved in different steps of physiological and pathological angiogenic processes, such as proliferation, maturation and survival of new blood vessels .Ang-2, which is part of a family of vascular growth factors that play a role in embryonic and postnatal angiogenesis and is involved in controlling microvascular permeability, vasodilation, and vasoconstriction by signaling the smooth muscle cells surrounding vessels , 7. In aGM-CSF is a cytokine secreted by macrophages, T cells, mast cells, NK cells, endothelial cells and fibroblasts and it functions as a white blood cell growth factor. Additionally, GM-CSF plays a vital role in angiogenesis through the regulation of VEGF and sVEGFR-1 . PDGF-BBThe aim of this study was to analysis serum concentrations of these markers associated with angiogenic processes, rarely investigated simultaneously in patients with IPF and sarcoidosis; looking for any possible differences and correlations with the main clinical parameters.Forty three non-smoking patients with sarcoidosis and 17 non-smoking patients with IPF characterized in our previous report  were enrSeventeen patients with IPF were diagnosed according to the ATS criteria  .Serum concentrations of Ang-2,follistatin, GM-CSF, IL-8, PDGF-BB, PECAM-1 and VEGF were measured in duplicate  using commercially available multiplex bead-based sandwich immunoassay kits . All assays were performed according to the manufacturer’s instructions. The system allowed simultaneous identification of 9 different molecules in a 96-well filter plate. We used 9 different sets of fluorescent dyed beads with capture monoclonal antibodies specific for each molecule to be analyzed. The appropriate molecule standards (50 μl/well) and samples diluted (50 μl/well) in plasma diluents were added to a 96-well filter plate and incubated for 30 min. at room temperature. After 3 washes, premixed streptavidin/phycoerythrin was added to each well and incubated for 10 min. followed by 3 additional washes. The beads were resuspended with 125 μl of assay buffer, and the molecule reaction mixture was quantified via the Bio-Plex protein array reader.The data analysis was performed using the Bio-Plex Manager software version 4.1.1. (Bio-Rad Laboratories). Values with a coefficient of variation over 12 % were discarded before the final data analysis. The concentration (pg/mL) of different analytes in the plasma samples was identified using standard curves from the multiplex assays. The minimum level of detection for VEGF was 3.27, 11.32 for Ang-2, 5.15 for PDGF-BB, 33.29 for PECAM-1, 8.41 for follistatin, 6.66 for G-CSF and 1.6 for IL-8.p < 0.05.The statistical analysis was conducted using Statistica 6.0 software . The normal data distribution was assessed using Shapiro-Wilk test. Homogeneity of variance was evaluated using Levene’s test. Comparisons between the study groups were performed using the ANOVA, Mann–Whitney or Kruskal-Wallis test, if the distribution of data was abnormal. Correlations were analyzed by Spearman’s test. All results were statistically significant if r = 0.62, P < 0.01), follistatin and PECAM-1  and PDGF-BB and VEGF .In the IPF patients in comparison with the control group and the sarcoid patients we observed significantly higher serum concentrations of GM-CSF, follistatin, PECAM-1 and IL-8. Serum concentrations of PDGF-BB were also significantly higher in the IPF patients than in the sarcoid patients but not than in the controls. In the IPF patients, serum Ang-2 and VEGF concentrations did not differ significantly from these obtained in the sarcoid group and the controls Table , and 7. r = 0.62, P < 0.0001), follistatin and PECAM-1  as well as PDGF-BB and PECAM-1 .In the sarcoid patients concentrations of serum Ang-2, follistatin, GM-CSF, IL-8, PDGF-BB, PECAM-1 and VEGF were similar to the values noticed in the control group. In patients with progressive and non-active  sarcoidosis the serum concentrations of all molecules were similar. No significant differences were observed between stage I, II and III-only serum Ang-2 concentrations in stage I patients was significantly higher than in stage II and III Table . In the Both in the IPF group and the sarcoid group no significant correlations between concentrations of the examined molecules and the pulmonary function parameters  Table .In our study elevated serum concentrations of IL-8, GM-CSF, follistatin, PDGF-BB and PECAM-1 in the IPF patients, but not in the sarcoidosis patients even in active progressive disease have been observed. Surprisingly, VEGF and Ang-2 concentrations  both in the sarcoid and the IPF patients were similar to the concentrations observed in the controls. We did not find any differences in serum concentrations of the examined cytokines obtained from patients with progressive i.e., active sarcoidosis and stable non-active sarcoidosis. To the best of our knowledge we were the first to demonstrate the serum concentrations of Ang-2, PECAM-1 and follistatin in IPF and sarcoidosis.Keane et al. , 19 demoNevertheless, Sekiya et al.  demonstrIn patients with sarcoidosis we previously observed significantly higher concentrations of GM-CSF in the BAL fluid from patients with active sarcoidosis in comparison with the control group . RecentlWe noticed significant elevation of PDGF in serum of IPF patients. Cao et al.  demonstrWe observed an increased serum concentration of follistatin in IPF, but not in sarcoidosis. Follistatin is effective in treating bleomycin -induced fibrosis by blocking the actions of activin and TGF-beta . RecentlIn our study serum concentrations of ANG-2 were also similar in IPF, sarcoidosis and the controls. Serum Ang-2 concentrations in stage I was higher than in stage II and III patients with saroidosis. ANG-2 is an essential factor for the vascular formation and is normally expressed in the endothelial remodeling tissues, while it is higher expressed during hypoxic processes . RecentlThe low angiogenetic serum activity in our patients with sarcoidosis may be, at least in part, explained by the findings of Kambouchner et al. . These aWe are not able to replicate some previously published results of circulating cytokine alterations. Methodological differences may account for our different, yet not contradictory findings: whereas many studies have employed single-cytokine ELISA tests, used plasma or culture supernatant samples, or had cohorts of newly diagnosed disease, we employed a multiplex assay, used serum samples, and had a cohort of chronic disease.Our study has limitations. A relatively small number of patients, especially with IPF, was included into the study and cytokines examinations were performed only in serum, but not in BAL. The results of Diffusing Capacity of the Lungs for Carbon Monoxide (DLCO) were available only in few patients as no correlations were estimated between DLCO and cytokines concentrations. No relationships were checked between the examined angiogenic molecules and other potential markers of sarcoidosis or IPF activity, for example: ACE (angiotensin converting enzyme) or LDH (lactate dehydrogenase), as no such data were available. It would be interesting to compare results obtained from patients with IPF with those at stage IV of sarcoidosis i.e., with fibrotic lung changes, it was very difficult however to collect a sufficient number of previously untreated sarcoid patients in IV stage.These results indicate that differences may exist in angiogenic activity between patients with parenchymal lung diseases. In contrast to sarcoidosis, IPF is characterized by a higher serum concentration of different molecules involved in the angiogenic processes.


Atomic level engineering of graphene-based materials is in high demand to enable customize structures and properties for different applications. Unzipping of the graphene plane is a potential means to this end, but uncontrollable damage of the two-dimensional crystalline framework during harsh unzipping reaction has remained a key challenge. Here we present heteroatom dopant-specific unzipping of carbon nanotubes as a reliable and controllable route to customized intact crystalline graphene-based nanostructures. Substitutional pyridinic nitrogen dopant sites at carbon nanotubes can selectively initiate the unzipping of graphene side walls at a relatively low electrochemical potential (0.6 V). The resultant nanostructures consisting of unzipped graphene nanoribbons wrapping around carbon nanotube cores maintain the intact two-dimensional crystallinity with well-defined atomic configuration at the unzipped edges. Large surface area and robust electrical connectivity of the synergistic nanostructure demonstrate ultrahigh-power supercapacitor performance, which can serve for AC filtering with the record high rate capability of −85° of phase angle at 120 Hz. Atomic level engineering of graphene-based materials is highly demanded for the customized structures and properties. Here, the authors show heteroatom dopant-specific unzipping of carbon nanotubes as a reliable and controllable route to customized 'intact crystalline' graphene-based nanostructures. A better controllability over unzipping mechanism is highly demanded for the minimal damage to the genuine graphene-based carbon structures16242526Unzipping of graphene basal plane is a valuable route to tailor carbon nanostructures123456781011121322 hybridized carbon framework, enable atomic-scale site-selective unzipping reaction. Although pristine CNTs are stable up to the electrochemical potential above 0.8 V, nitrogen-doped CNTs (NCNTs) are readily unzipped below 0.6 V. Detailed investigation on the reaction mechanism reveals that substitutional pyridinic nitrogen (Np)-dopant can specifically initiate CNT wall unzipping. Such a dopant-specific unzipping at moderate potential enables fine controllability of unzipping level and intact crystallinity of the unzipped structures with well-defined edge configuration. Taking advantage of these unique features, we are able to synthesize intact crystalline graphene-based nanostructures, where unzipped graphene nanoribbons are seamlessly connected to the CNT strands. This structural feature offers synergistic properties comprising large surface area and robust electrical conductivity, which are highly desirable for electrochemical applications, such as energy storage. As a representative application, we demonstrate ultrahigh-power double-layer capacitors (DLCs) for alternating current (AC) line-filtering performance.In this work, we present dopant-specific electrochemical unzipping of CNTs as a controllable method for intact crystalline unzipping. Heteroatom dopants, such as nitrogen substitutionally incorporated into sp2SO4 aqueous solution at a positive potential of 0.6–0.8 V, electrochemically developed high-energy oxygen species react with the carbon atoms nearby the Np-dopant sites and initiate unzipping242SO4 solution and subsequently unzipped (unzipped . Eventuaunzipped  are formunzipped .29ID/IG) from 1.06 to 1.52 and up-shift of G band position from 1,581 to 1,588 cm−1 result from the decrease of in-plane sp2 domain size upon C=C bond breaking reactions263132Chemical structure of typical unzipped materials was characterized by X-ray photoelectron spectroscopy (XPS) and Raman spectroscopy. Deconvoluted C1s XPS spectrum  shows thp peak is specifically reduced during unzipping, whereas quaternary nitrogen (NQ) is not influenced. Concurrently, Fe2p XPS results (p atoms to generate FeN4 moieties in the graphene plane+) can replace the Fe2+ ion to form H2N4 moiety  is easily tunable by adjusting NH3 gas partial pressure during the synthesis of NCNTs36p-dopant-specific unzipping is triggered at the significantly reduced potential of 0.6 V  enables the fine controllability of unzipping propagation. Ic) is another measure to monitor the unzipping progress, particularly in terms of surface area enhancementIc rapidly increases in the early stage because of the enlargement of surface area by gradual exposure of the inner surface of NCNTs, and the rate enhancement of Ic slowdowns after 4 h  were measured at 1–400 V s−1 . Reproducan rate . This facan rate Electrochemical impedance spectroscopy further confirms the outstanding power performance of the DLCs with unzipped structures. 4142434445In a DLC, series resistance, which hampers the power capability of the devices, can be simply considered as the sum of electronic and ionic resistance. The vertically aligned unzipped morphology of our electrode materials can effectively minimize the resistive elements. The vertical alignment offers idealized straight pathway for facile ion transport and homogeneous distribution of stored charges at electrode surfaces4644CA, F cm−2) is also calculated  of DLCs fabricated with 5-μm-NCNT forest, 5- and 1-μm-unzipped structures are 616, 468 and 212 μs with 2.68, 1.24 and 1.15 Ω of total resistance at 120 Hz, respectively. These values are sufficiently smaller than the standard 8.3 ms period, which is crucial for the effective AC line-filtering at 120 Hz. The unzipped nanostructures also demonstrate a very small relaxation time constant  at 120 Hz, significantly higher than previous carbon based-electrodes. This CV value is approximately three orders of magnitude higher than those of typical AECs (∼10−4 F cm−3)4344In this work, we report the ultrahigh-power performance of unzipped structure-based DLCs. In contrast to the intensive research efforts for high-energy-density supercapacitors155253542-hybridized carbon plane allows the transformation of structural dimension among zero-, one-, two- and three-dimensions. Overall, customized dopant-specific unzipping of graphene-based materials possibly guides us to rarely explored opportunity for multi-dimensional material design with atomic level precision as well as physicochemical property tuning, which are generally desired for energy storage, environmental remediation, nanomedicines and other versatile applications. Moreover, this precisely controllable unzipping mechanism grants a significant step forward to idealized intact crystalline graphene nanostructures in conjunction with the potential advances in the precise control of dopant position, dopant density and the chirality of CNTs.As demonstrated from the record-high power DLCs, delicate manipulation of graphene-based nanostructures may offer straightforward routes to high-performance applications. The highly specific reaction features of our unzipping mechanism can serve as a generic principle to diversify graphene-based nanostructures, whose physicochemical properties are strongly dependent on their structural dimension and defect formation123456783 gas partial pressure and plasma power (2/Si was floated on diluted hydrofluoric acid (HF) solution to separate NCNT forests from SiO2/Si substrate by wet-etching of SiO2 layer. The floated NCNT forests were carefully transferred onto a glassy carbon electrode. To enhance the mechanical and electrical contact between NCNT forests and glassy carbon electrode, the recovered electrode was thermally annealed at 65 °C for 12 h. The NCNT electrode was used as working electrode in typical three electrode configuration with mercury/mercury (I) sulfate reference electrode and platinum wire (counter electrode). The unzipping was carried out at room temperature in 1 M H2SO4 solution with positive electrical potential from 0.5 to 1.0 V for various unzipping period. Before each unzipping reaction, the electrolyte was purged with high purity N2 gas over 10 min.NCNTs were grown by plasma-enhanced chemical vapour deposition with Fe catalyst. The density of N dopants was easily controllable by adjusting NHma power . As-grow2/Si substrate. Raman and XPS spectra were recorded with ARAMIS (Horiba Jobin Yvon) and Sigma probe (Thermo VG Scienfitic), respectively.HR-TEM images were collected on a double-aberration corrected HR-TEM  located at KAIST Analysis Center for Research Advancement, operating with a monochromator (excitation 0.06) at 80 kV accelerating voltage. The structural changes were also measured by field-emission scanning electron microscope . Samples were prepared by critical point drying to maintain the as-synthesized vertically aligned structure. AFM images were obtained with scanning probe microscope . Samples were prepared by drop-casting from ethanol suspensions onto SiO−2) on Pt foil were used as an electrode for ultrahigh-power DLCs. Before DLC assembly, the unzipped structures were electrochemically reduced at −0.6 V for 30 min to remove dispensable oxygen functional groups. The prototype DLCs were composed of two identical unzipped nanostructure electrodes and separator. All electrochemical characterization was carried out with a potentiostat  using 1 M KOH solution as an electrolyte under ambient environment. All CVs in CA, F cm-2)Unzipped nanostructures ; Z"(f) is the imaginary part of the impedance.where CV, F cm−3) was calculated with equation (2)The frequency specific volumetric capacitances  and C"(f),The specific capacitance of DLCs prepared with unzipped nanostructures could be calculated by using C′(f) is the real part of specific capacitance; C"(f) is the imaginary part of specific capacitance; Z′(f) and Z"(f) are real and imaginary parts of the impedance; |Z(f)| is the absolute value of the impedance.where Ab-initio Simulation Package codes. The Perdew–Burke–Emzerhof exchange-correlation energy functional and projected-augmented wave potentials were employed. A plane-wave kinetic energy cutoff of 400 eV and 1 × 1 × 1 gamma k-point were used for 12 unit cells of  single-walled CNT with 240 atoms. The supercell size is 20 × 29.514 × 20 Å3 with the tube axis along the y direction, and the diameter of model is about 6.726 Å. Atomic forces were relaxed <0.025 eV Å−1.To understand the mechanism for electrochemical unzipping of CNTs, first-principles DFT calculation was performed using Vienna How to cite this article: Lim, J. et al. Dopant-specific unzipping of carbon nanotubes for intact crystalline graphene nanostructures. Nat. Commun. 7:10364 doi: 10.1038/ncomms10364 (2016).Supplementary Figures 1-17, Supplementary Table 1, Supplementary Note 1 and Supplementary References.


Graphene is a highly versatile two-dimensional material platform that offers exceptional optical and electrical properties. Of these, its dynamic conductivity and low effective carrier mass are of particular interest for optoelectronic applications as they underpin the material’s broadband nonlinear optical absorption and ultra-fast carrier mobility, respectively. In this paper, we utilize these phenomena to demonstrate a high-speed, in-fibre optical modulator developed on a side-polished optical fibre platform. An especially low insertion loss (<1 dB) was achieved by polishing the fibre to a near atomically smooth surface (<1 nm RMS), which minimized scattering and ensured excellent contact between the graphene film and the fibre. In order to enhance the light-matter interaction, the graphene film is coated with a high index polyvinyl butyral layer, which has the added advantage of acting as a barrier to the surrounding environment. Using this innovative approach, we have fabricated a robust and stable all-fibre device with an extinction ratio as high as 9 dB and operation bandwidth of 0.5 THz. These results represent a key step towards the integration of low-dimensional materials within standard telecoms networks. Graphene is the most prominent of an emerging catalogue of two-dimensional materials that are currently spear-heading intense research across the optoelectronics community23710An alternative approach that offers a straightforward route to ultra-fast modulation is to exploit the strong saturation of graphene’s absorption due to Pauli blocking, whereby the generated carriers fill up the valence bands preventing any further transitions. By employing a pump-probe scheme, all-optical modulation has recently been demonstrated in a graphene-coated microfibre device with an operation speed approaching 200 GHz1013In this paper, we present a robust, all-optical modulator with a large 9 dB extinction ratio and an operation bandwidth of 0.5 THz. The device is built into a side-polished window of a standard optical fibre and is thus immediately compatible with existing fibre networks. By retaining a small cladding buffer between the core and the polished window, the insertion loss of the device is less than 1 dB, which represents a record low value for this type of device1516−1 full width half maximum (FWHM) of the 2D peakSchematic representations of our graphene-coated modulator are illustrated in To illustrate the principle of the enhanced light-matter interaction, the longitudinal cross-section of the device is shown in We start our investigations of the transmission properties of the graphene-PVB device by comparing the losses of the TM and TE modes using a polarized continuous wave (CW) laser at 1550 nm. The input power was set to 0 dBm and the polarization was tuned between the modes using a half wave plate. When coupling into the TE mode of the device, the reduction in output power was <1 dB, which is commensurate with insertion losses of commercially available fibre devices. The experiment was repeated again for the TM mode and the power was at a minimum of −31 dBm, as expected for our CVD-produced graphene film. This large, 29 dB attenuation difference between the TM and TE modes provides a good indication of the high purity of our graphene sheet and the strong light-matter interaction that has been achieved. We note that the PVB material used here is largely transparent across the transmission window of the silica fibre and thus the device should be operable across the entire telecom band. To test the broadband nature of the device, additional transmission measurements were undertaken at wavelengths spanning 1425–1600 nm, from which it was confirmed that the extinction ratio did not drop below 25 dB in this region see .ave~10 mW the absorption of the TM mode is linear, however, for higher input powers the absorption becomes nonlinear and the transmitted light starts to increase exponentially, eventually reaching a transmittance of 60% for Pave~32 mW. In contrast, the transmission of the TE mode remains linear when the input power is increased up to the maximum available power. This is due to the minimal interaction between the TE mode and the graphene sheet; a phenomenon that forms the basis of the graphene polarizerCharacterization of the nonlinear saturable absorption induced by Pauli blocking was then undertaken using a high power fibre laser operating at 1540 nm, with a duration of 750 fs (FWHM) and a repetition rate of 40 MHz. ave~20 mW, corresponding to a peak power of Pp~720 W) and a weak probe component . The probe was modulated at 100 Hz using an optical chopper connected to a lock-in amplifier to discriminate between the two signals. Polarization controllers were used to ensure that both the pump and probe were aligned to couple into the TM surface wave, and a 10 ps adjustable delay was inserted into the probe’s path to control the overlap between the two pulses. The pump and probe pulses were then recombined using a fused tapered coupler before being connected to the input arm of the device under test. 1~0.1 ps and τ2~0.4 ps, in good agreement with previous reports2 has been found to be directly proportional to the crystal coherence length in the graphene sheet and thus a possible route to increasing the modulator’s intrinsic speed would be to engineer a level of disorder into the lattice.In order to determine the temporal dynamics of the saturable absorption we employed a simple pump-probe modulation scheme, as illustrated in In conclusion, we have designed and experimentally demonstrated a graphene-based all-optical modulator with a high extinction ratio of 9 dB and a speed of 0.5 THz. When compared to previous devices, our method results in an enhanced interaction with the graphene sheet to produce a significant increase in modulation depth, while at the same time ensuring a large decrease in insertion loss. This is best illustrated by applying the figure of merit (FOM) proposed in ref. Standard Corning SMF-28 step-index single-mode fibres with 125 μm diameter cladding, 8.2 μm diameter core, and 0.14 numerical aperture are used for the construction of the devices. An aluminium block was prepared with large-radius curved-sides, a 1 cm long flat-centre and U-shape cross-section groove cut down the middle. The radius of the two curved-side sections was fixed at 30 cm to form an adiabatic transition region and the depth of the groove was ~65 μm to allow polishing to the desired depth. A 3 cm long section of the fibre was stripped of its buffer jacket and bonded into the groove with a wax. The bulk of the top section of the fibre was removed using 3 μm aluminium oxide film, following which it was polished to a surface quality of~1 nm RMS using a proprietary polishing film (UltraTec).2/Ar gas. After that, the temperature was increased to 1000 °C with 6% H2/Ar gas flow of 100 sccm and the chamber pressure maintained at 20 mBar. The Cu foil was annealed at these conditions for 30 min. The graphene growth was performed by flowing 5% CH4/Ar together with 6% H2/Ar gases for 20 min with flow rates of 100 sccm and 210 sccm, respectively. Finally, the CH4/Ar flow was turned off to finish the graphene deposition and the Cu foil was cooled down naturally.A typical CVD system for the fabrication of graphene is shown in the −1 to 2850 cm−1 with a resolution of 0.1 cm−1. The Raman features of mono-layer graphene are the G peak at ~1580 cm−1 and a symmetric 2D peak at ~2700 cm−1 with full-width half maximum (FWHM) of~25 cm−1. The negligible D peak, normally seen in the spectrum near ~1350 cm−1, indicates that high quality  graphene has been produced. In general, the appearance of the D peak indicates the disorder in the carbon lattice such as the domain boundaries, lattice defects or distortion. In addition to the shape of the 2D peak, it is also known that the ratio of I2D/IG can be used to determine the number of graphene layers. The typical I2D/IG ratios of single layer and bilayer exfoliated graphene are ~2–3 and slightly less than 1, respectively. For our as-grown CVD graphene layer shown in The as-grown mono-layer graphene has dimensions of 80 mm × 90 mm, as shown in The optical transmission loss of the device was measured with a 1550 nm diode and the cut-back technique was used to remove extraneous losses. Measurements were undertaken as a function of wavelength using a low power continuous wave Tunics laser as the source and a Newport 818-IR detector. The source used for the high peak power experiments was a OneFive Origami with a bespoke pulse duration of 750 fs FWHM. The carrier dynamics were estimated by comparing the modulated response directly with the pump pulse. As the pump is symmetric, the asymmetry in the tail is attributed to the slow recovery of the carriers. The relaxation times were then determined by fitting the extended tail with a convolution of 2 exponentials.Numerical simulations were undertaken using the COMSOL Multiphysics with the RF module and the LiveLink to MatLab.How to cite this article: Zhang, H. et al. Enhanced all-optical modulation in a graphene-coated fibre with low insertion loss. Sci. Rep.6, 23512; doi: 10.1038/srep23512 (2016).


The questions’ judged answerability level on the two scales was highly correlated. For both scales, consensus questions were rated more answerable than the non-consensus questions, with illusion questions falling in-between. The result for the illusion questions indicates that a feeling of answerability can be created even when it is unlikely that somebody can come up with an answer. The results also showed that individual difference variables influenced the answerability judgments. Higher levels of belief in certainty of knowledge, mankind’s knowledge, and mankind’s efficacy were related to judging the non-consensus questions as more answerable. Participants rating the illusion questions as answerable rated the other answerability questions as more, or equally, answerable compared to the other participants and showed tendencies to prefer a combination of more epistemic default processing and less intellectual processing.Occasionally people may attempt to judge whether a question can be answered today, or if not, if it can be answered in the future. For example, a person may consider whether enough is known about the dangers of living close to a nuclear plant, or to a major electricity cable, for them to be willing to do so, and state-authorities may consider whether questions about the dangers of new technologies have been answered, or in a reasonable future can be, for them to be willing to invest money in research aiming develop such technologies. A total of 476 participants, for each of 22 knowledge questions, either judged whether it was answerable today (current answerability), or judged when it could be answered (future answerability). The knowledge questions varied with respect to the expected consensus concerning their answerability:  In daily life and in science people are sometimes faced with questions that they do not immediately know the answer to. Examples are questions such as “Is it safe to use a cell-phone daily?”, “Can this person be considered guilty beyond a reasonable doubt?”, or “Does this medicine have serious side effects?” Moreover, if a person thinks that a question currently has not been answered, they may consider how long it will take before it will be answered, or if it can be answered at all. For other questions, people may think they know the answer, only to find out that their judgment was premature. Many tragic human catastrophes may, at least in part, have occurred because people misjudged the answerability of important questions, e.g., the Thalidomide catastrophe in the 1950s.answer to mean an answer that is correct and that is provided with good arguments at a relevant level of specification. In this context there are thorny philosophical issues related to, for example, the concepts of truth and knowledge that have been debated by philosophers for centuries. However, this paper does not try to solve such issues. Thus, for example, at a philosophical level, it may presently not be possible to convincingly claim that there is an absolute difference between answerable and non-answerable questions. In spite of this, we note that people, both in science and in everyday life in fact make judgments about when in the future, questions judged not to have been answered, are likely to be so, if ever. The present study investigated people’s subjective judgments of questions’ answerability.The notion that questions can have correct answers is commonly taken for granted in everyday life and in science. In the present research, we use the term Such judgments are important because they may influence other judgments and decisions relevant in everyday life, for example decisions whether to allow new techniques or substances in specific contexts such as in medicines, clothes, foods, or building materials. An example of a future answerability judgment is how many test trials should be made before we can be sure that a new drug is not dangerous to the public? Other contexts where answerability judgments are made in everyday life, including work life, concern allocation of resources to research project applications or to public and other inquiries. For example, if it is reasonable to think that a research project can answer its questions during the time the research money is applied for? As far as we know no research has studied answerability judgments in the broad sense discussed here.Much of previous research on question answerability judgments has been limited to questions where people tend to agree on the answer and has tended to study isolated aspects of the broader answerability question, for example, if I myself or someone else knows the answer to a posed question. Thus, studies of “I know/don’t know” judgments have been made in different fields of psychology. To illustrate, research in forensic psychology has explored participants’ ability to separate out questions on “information not seen” in a video clip e.g., , in geneothers’ knowledge. To illustrate, Research on people’s confidence in their answers to different types of memory questions has in many contexts found a tendency for people to show overconfidence in their responses  have been addressed in educational psychology orientated toward science communication e.g.,  and in rEvidence in risk research shows that differences in cultural values such as hierarchical/egalitarian and individualistic/communitarian may shape individuals’ beliefs about the existence of scientific consensus on such topics as climate change and disposal of nuclear waste . Other rignorance, taken in a broad sense. Researchers have also presented taxonomies of ignorance  and with respect to question answerability (answerability consensus). There are at least three types of possible answerability consensus: whether there is some answer to the question today, whether it can be answered in the future, and about the answerability of specific types of questions. A common conclusion is that people have a tendency to trust socially prevalent understanding e.g., . SociallIn this research we investigated answerability judgments of three types of questions: questions for which we expected a high degree of consensus regarding their answerability (consensus questions), questions for which we expected a low degree of such answerability consensus (non-consensus questions), and questions that may appear answerable but where some information necessary to answer it was missing (illusion questions). However, we do not claim that there is an absolute qualitative difference between these question types, rather the difference may be a question of degree. Consensus and non-consensus questions were used in this research because it is of interest to study the consistency in people’s answerability judgments of questions with different degrees of expected answerability. In addition, we were also interested in studying the degree to which individual difference variables influenced questions with different levels of expected consensus about their answerability (further elaborated below). The illusion questions were included in order to examine the extent to which missing information in the questions might be compensated for by the use of other types of information such as conceptions about the answerability of specific types of questions.We expected that consensus questions would be rated higher in answerability than non-consensus questions (Hypothesis 1). This was partly because we expected that it would be relatively easy for the participants to either provide what they thought was the correct answer to the question (thus showing that is was answerable), or to imagine some easily performed way to get the answer to the question. The illusion questions belonged to geometry and physics and, since we had purposefully eliminated information necessary to answer them, they show similarity to the questions used in the witness psychology and memory studies reviewed above. Moreover, they were designed to appear fairly elementary and computable and we believed many participants (possibly from their school experience) would think that the answer to such elementary computational questions in general is possible to compute. This is in line with current answerability of the question and the other scale to when, if ever, a question can be answered. These two scales were included because it is reasonable to think they are commonly used, explicitly or implicitly, when people judge the answerability of questions in everyday life. In addition, we were interested to compare the answerability levels of the judged questions on the two scales since if the rank order in answerability is stable between the two scales this is an indication of some stability in the processes generating the answerability judgments.Answerability judgments can be made on different types of scales and it is of interest to understand the extent to which the level of the answerability judgment varies as an effect of the used scale. Therefore we compared two kinds of answerability scales with respect to their effect on the level of answerability judgments. One scale related to the per se, may be irrelevant to their realism. Therefore, we also investigated the relation between answerability judgments and measures of individuals’ cognitive and personality properties, henceforth called individual difference variables, that it seems reasonable to believe will be associated with the level of the answerability judgments. In general it would seem that consensus type questions are more likely to be judged by use of quick and fairly automated processes of a System 1 kind  answers may be out of human reach or context dependent . We expeBeliefs about the extent and usefulness of mankind’s knowledge as such also illustrate belief about epistemic issues. Given that more knowledge might be expected to be associated with greater possibility to answer knowledge questions, we hypothesized that belief in a larger body of mankind’s knowledge would be correlated with higher answerability judgments (Hypothesis 4).A further type of belief concerns beliefs about mankind’s effectiveness to reach, for example, epistemic goals , that is, mankind’s efficacy. We expected that participants who believed more in mankind’s efficacy would tend to judge questions as more answerable (Hypothesis 5).EPI-r scale  and preference for a deeper intellectual type of processing measured by EPI-r Intellectual (similar to system 2 processing). We expected that participants with default processing preferences would tend to choose interpretations that come quickly to mind and are easy to handle and therefore would find questions more answerable. Furthermore, we expected that preference for intellectual processing would be associated with lower answerability judgments since participants high in this preference would problematize the possible constructions of answers to questions more than participants low in this preference (Hypothesis 6). Our reasoning here was that people with a higher default processing preference may be more prone to rely on general rules of thumb , while intellectual processing may make it easier to notice that crucial information needed to answer the question may be missing.Shallow or deep processing might influence answerability judgments. Two kinds of processing preferences are measured by the -r scale : prefereOptimism may also affect the level of answerability judgments. People with generalized personal optimism tend to interpret things in a positive way and are less likely to give up . As optiIn total, 476 participants completed the 22 answerability questions in one of the two answerability scale variants . The mean age was 28 years (range 18–78 years). Of the 476 participants, 236  completed the answerability questionnaire using the current answerability scale. The remaining 240 participants  used the future answerability scale. As a reimbursement, participants participated in a lottery for a cinema ticket.The present study followed ethical guidelines in Sweden for survey data. Participants were recruited from a pool of adults that had already actively volunteered and signed up for participation in psychological research and can thus be considered consciously aware of participation in general. They were provided with information about the purpose of the study via e-mail and were told that participation was not mandatory. In the email they were informed that their answers only would be used for research purposes, that they could withdraw at any time. The email also provided relevant contact information. Participants gave their consent by clicking on a survey link. When clicking on the link for participation participants were randomized to four groups. Approximately half of the participants were randomized to each of two scale variants: the current answerability scale and the future answerability scale (described below). In order to control for any ordering effects within each scale variant group the order of the question blocks was altered, constituting two order conditions. In the first order condition (the belief in certainty of knowledge-first condition) the participants answered the belief in certainty of knowledge items first, then the answerability questions, and then three other measures-the LOT-r, EPI-r, and mankind’s efficacy, in a randomized order. In the second order condition (the belief in certainty of knowledge items-last condition) the participants answered the questions in the following order: first the answerability questions, then the three other measures-the LOT-r, EPI-r, and mankind’s efficacy-in a randomized order, and last the belief in certainty of knowledge items. Finally, two questions about mankind’s knowledge and background questions were answered.1 participants answered all the 22 answerability questions and data from that set of people were used for further analysis. Thus, the response rate was 33%.In total 1462 individuals from participant pools at the University of Gothenburg were invited to answer a web-questionnaire. It was not possible to go back to previous pages in the questionnaire to change answers. If participants left a question unanswered, they were kindly reminded but not forced, to complete the question. This rendered 639 answers. Out of these 639, 476Appendix 1). However, at the same time we kept the total number of questions reasonably low in order to achieve a good response rate and an even answer quality throughout the questionnaire . The remaining two questions were illusion questions in which a crucial detail necessary to compute the answer was missing. For example, in the question “How large is the area of an ellipse with a minor axis of 2 cm?” the length of the major axis was absent. Three types of questions were used. There were eight F = 23.45, Bonferroni correction (p < 0.01) which supported the researchers’ intuitions. However, the illusion questions did not differ significantly from the consensus (p = 0.14) nor the non-consensus questions, (p = 0.22).In a pre-study 100 participants from a student pool at the University of Gothenburg rated various aspects of the 22 question-items. Participants estimated the proportion of adult Swedes that would agree that the question is answerable . On average participants judged that 74% of the Swedes would agree that the consensus questions were answerable, but estimated that only 55% of the Swedish population would agree that the non-consensus questions were answerable. Participants further judged 64% of the Swedes would reckon the illusion questions answerable. The difference between consensus and non-consensus questions was significant The questions in the present study were organized in pairs so that questions of different categories were matched with each other. There were two pairs of questions containing one illusion and one consensus questions, six pairs containing one consensus and one non-consensus question, and three pairs containing two non-consensus questions. The just described pairs of answerability questions were presented in an order randomized for each participant.answerability means that the question can be answered on a level that is exact and relevant enough. E.g., for the question “how tall is the world’s smallest living creature ever”, the answer “less than a meter” is not exact enough to be counted as correct.” The instructions are further described in Appendix 2.For both scale variants , the instructions stressed that the judgment task was not to provide the answer as such to the question but to judge the answerability of the question. Moreover, the participants were instructed that “Each screen in the questionnaire presented two questions for which the participants were to give answerability judgments. On the top of each screen was a reminder formulated as follows: “Reminder: for the question to be judged as answerable it is necessary:” (the next three sentences each appeared on its own line) “That at least one person in the world can answer the question. That the answer can be answered correctly and that good arguments for the answer can be provided. That the question can be answered in a sufficiently exact and relevant way.”Figure 1.Two scales were prepared for the answerability judgments. One of the scales, “the current answerability scale” asked the participant to judge the probability that the question could be answered by at least one person now living in the world. The other scale, “the future answerability scale”, more openly directed the judgment to when the question considered can (if ever) be answered. The two scales are shown in If scientists try hard enough they can find the truth to almost everything” and “Scientists can ultimately get to the truth”. Higher values indicate higher belief in certainty of knowledge. The items were rated on a five-point scale, ranging from 1 = Do not agree at all to 5 = Totally agree.Two items from a factor tapping belief in certainty of knowledge in the epistemic beliefs measure presented by Mankind can always manage to solve difficult problems if it tries hard enough”, “Even if hard times threaten mankind, mankind will find ways to reach its goals”, “In unexpected situations, mankind will find ways to act”, and “Even in unexpected situations I believe mankind can cope well”. Items were answered on a five-point scale ranging from 1 = Do not agree, to 5 = Totally agree. Cronbach’s alpha was 0.77.A measure was constructed in order to tap a belief in mankind’s ability to complete tasks and reach goals. The items were similar to self-efficacy items, but instead addressed mankind’s efficacy. The four items were: “How much does mankind know of all there is to know?” and “How much does mankind know of all that is important to know?” Items were answered on a scale ranging from 0 to 100% in intervals of 10. Cronbach’s alpha for the items was 0.66, which although not high can be considered acceptable for research purposes  and intellectual processing . Questions were answered on a five-point Likert-type scale, ranging from 1 = Do not agree to 5 = Agree completely. Cronbach’s alpha was 0.71 for default processing, and 0.81 for intellectual processing.The Epistemic Preference Indicator-Revised  measuresIn uncertain times, I usually expect the best”. The items were answered on a five-point scale ranging from 1 = Do not agree to 5 = Totally agree. Cronbach’s alpha was 0.78.Life orientation test revised measures the degree of optimism regarding oneself and has six items . An itemTable  Since the future answerability scale starts with two categories of a nominal type, these categories were recoded into the same category in order to make the scale more ordinal. Thus, after recoding 1 = I or someone else knows the answer to the question, 2 = Can be answered within 1 year, 3 = Can be answered within 10 years, etc. Medians and interquartile range were used for the future answerability scale.Means, medians, SDs, and interquartile range for the answerability judgments for the three types of questions are shown in Table 1, for the current answerability judgments, consensus questions were perceived to be most answerable (M = 93%), non-consensus questions the least answerable (M = 47%), and illusion questions were rated in between (M = 77%). Figure A1 in Appendix 3 shows that the answerability questions’ mean ratings ranged from almost 100% answerable to approximately 15% answerable.As can be seen in Mdn = 1, “I or someone else knows the answer to the question”) and the non-consensus questions least . Most of the respondents (75th percentile) rated the non-consensus questions to be answered within a maximum of 50 years. The response alternative Can never be answered was chosen 12% of the time.For the future answerability judgments consensus questions and illusion questions were rated most answerable .2 The six individual differences variables were entered as covariates to control for their influence .In order to investigate differences between consensus and non-consensus questions a mixed ANCOVA was conducted with the within-subject factor of question type (consensus vs. non-consensus) and, to control for ordering effects, the between-subject factor F = 35.41, p < 0.001, partial η2 = 0.15. We found no effect of the order factor , F = 0.01, p = 0.942. No interaction effect was found between the question type and order factors, F = 0.25, p = 0.618.Consensus questions were rated as significantly more answerable than non-consensus questions, PS) was used, following PS estimates the probability that a score randomly drawn from population a will be greater than a score randomly drawn from population b.Due to the ordinal properties of the future answerability scale non-parametric tests were used. To estimate the effect sizes the probability of superiority estimator (U-test showed that there were no differences in answerability judgments for the consensus questions as a consequence of whether the belief in certainty of knowledge items were rated before or after the answerability questions (p = 0.957). However, there was a significant difference for non-consensus questions due to rating belief in certainty of knowledge before or after the answerability judgments, U = 8484.50, z = -2.494, p = 0.013. PS = 0.59. Therefore the results for the future answerability of the non-consensus questions were analyzed jointly and separately per order condition and reported when relevant.A Mann–Whitney z = 10.24, p < 0.001, PSdep = 0.74.To investigate differences between future answerability for the consensus and non-consensus questions a Wilcoxon signed rank test was used. The consensus questions were rated as being possible to answer in a nearer future than the non-consensus questions, sr(22) = -0.48, p < 0.03. Thus, on average, the questions considered unlikely to be answered today were considered to be answered in a more distant future. .The median of each question item on the current answerability scale was correlated with the median of each corresponding question item on the future answerability scale. This correlation was significant and very high, Table The means and SDs for the individual difference variables and the current answerability ratings of consensus, non-consensus and illusion questions and Pearson correlations between these variables are shown in r(226) = 0.28, p < 0.001. Also, people who believed more in mankind’s efficacy rated the non-consensus questions as more answerable, r(216) = 0.22, p = 0.001.There were no significant correlations between the individual differences variables and the answerability ratings on the current answerability scale for the consensus questions. However for the non-consensus questions, people that rated a high belief in certainty of knowledge rated these questions as more answerable, F = 5.26, p < 0.001. The regression analysis showed that the individual difference variables explained 13% of the variance in the answerability judgments. Furthermore, belief in certainty of knowledge , mankind’s efficacy  and optimism  yielded significant β-values.In order to explore how much variance in the answerability ratings of the non-consensus questions was explained by the individual difference variables when the current answerability scale was used, a multiple regression with the dependent variable answerability of non-consensus questions and the independent variables belief in certainty of knowledge, EPI-Default, EPI-Intellectual, mankind’s efficacy, optimism and mankind’s knowledge was conducted. The result was significant r(216) = -014, p < 0.047.A negative correlation was found between the answerability ratings of the illusion questions and optimism, thus, the more optimistic people rated illusion questions as less answerable, One hundred and one participants in the current scale condition rated both of the illusion questions as 100% answerable, and were thus “tricked” by the illusion questions. Interestingly, on average these 101 participants also rated each of the other questions higher or equally high in answerability compared to other participants. In order to investigate if epistemic processing preference differed between the tricked and non-tricked participants, a general EPI-score was computed with the formula . Tricked participants did not score significantly higher on the EPI–score.Table The correlations between the future answerability scale and the individual difference measures can be found in p < 0.05) were found between the consensus questions and the individual difference measures for the future answerability scale. As can be seen in Table 3, rating the non-consensus questions to be more answerable in the near future was associated with higher ratings for mankind’s efficacy rs(214) = -0.15, p = 0.028, and mankind’s knowledge rs(214) = -0.15, p = 0.026.No significant correlations (rs(113) = -0.27, p = 0.004. Likewise, the correlation between mankind’s efficacy and the answerability values for the non-consensus questions was more pronounced when belief in certainty of knowledge was judged before answerability judgments, sr(113) = -0.23, p = 0.016.The correlation between mankind’s knowledge and the answerability values for the non-consensus questions was stronger when belief in certainty of knowledge was rated first, Table 3, rating the illusion questions as being more answerable in the near future was associated with higher ratings for mankind’s knowledge rs(214) = -0.14, p = 0.036, and lower EPI-intellectual rating, rs(214) = 0.19, p = 0.005. Thus, higher answerability ratings for the illusion questions were associated with lower values on the EPI-intellectual scale.As can be seen in n = 182) had a significantly lower EPI-score , t(432) = -2.20, p = 0.028, d = -2.20.In total 182 participants claimed the illusion questions could be answered today and were thus tricked by the illusion questions. Comparing the medians, tricked participants judged each of the other questions to be answered at the same time or sooner than non-tricked participants. The tricked participants (This study investigated judgments of current and future answerability of three types of knowledge questions: consensus questions (high level of expected consensus about their answerability), non-consensus questions (lower expected level of consensus about their answerability), and illusion questions . We also investigated whether there was a relationship between various individual difference variables and the level of the answerability judgments.Questions rated low on answerability on the current scale were also rated to be answerable further off in the future. The finding that the level of judged answerability was fairly strongly correlated between the two scales indicates that there is some stability in the processes generating answerability judgments. This result is also in line with the conclusion reached by The consensus questions were rated as having a high level of answerability and were, as expected, judged as more answerable than the non-consensus questions, independent of the scale used. A possible explanation for the high ratings of the consensus questions is that fairly automated memory processes more often quickly identified a correct answer or classified the consensus questions as being a type of question that could be known by myself or by others. This explanation coheres with research on “don’t know” judgments which shows that spontaneous quick processes may quickly contribute to the assessment of many questions e.g., . ResultsThe non-consensus questions were given the lowest answerability ratings of the three question types. In line with the considerations above, the answerability ratings for these questions may have been more dependent on elaborated and deliberative thinking, compared to the consensus questions. For example, it is likely that there was less experienced consensus about a possible answer to many of these questions and that for this reason the advice offered from socially prevalent knowledge compare  may haveper se. Interestingly, participants who failed to notice the missing information in the illusion questions also rated other questions either higher or equally high in answerability.Many participants rated the illusion questions as answerable today even though crucial information was missing. The results for the illusion questions indicate that a “feeling of others knowing” can be created even when it is unlikely that somebody can come up with an answer. The participants who were tricked by the illusion questions showed tendencies to have an epistemic preference for higher default processing in combination with lower intellectual processing. High answerability judgments for the illusion questions may therefore possibly be explained by shallow processing based on general beliefs , in contOverall, the individual difference variables all showed acceptable or satisfactory Cronbach’s alpha values and were moderately related to answerability judgments. In line with our hypothesis, when an effect was found it was related to the answerability of non-consensus questions, not to the consensus questions. However, it should be noted that the weaker correlations for the consensus questions may also be due to their lower variability, compared with that of the non-consensus questions and this issue should be further explored in future research.Together the individual difference variables explained 13% of the variance of the answerability judgments for non-consensus questions when the current scale was used. This can be seen as an irrelevant influence on the level of the answerability judgments in the sense that variance in individual difference variables is not usually considered likely to contribute to better quality in judgments, that is, more realistic values. Future research should investigate if there are specific levels of some individual difference variables that are beneficial for producing realistic answerability judgments. Generally speaking, in important applied contexts it may be worthwhile to develop approaches that can help to decrease the negative influence of individual difference variables.It is also noteworthy that the participants who believed more in certainty of knowledge also rated the more challenging questions (the non-consensus questions) as more answerable, especially for the current scale, providing some support for our hypothesis. They also tended to believe more in mankind’s knowledge  and especially to believe more in mankind’s efficacy with respect to solving difficult problems and troubles. These associations may also have contributed to the tendency for participants who believed more in mankind’s knowledge and efficacy to give higher answerability ratings. Future research should investigate whether people who believe more in certainty of knowledge and mankind’s efficacy differ from other people in the realism of their answerability judgments. For example, do such people, when they consider real-life complex issues, have a tendency to underestimate the number of variables relevant to consider or the complexity of the possible interactions between variables?When it comes to epistemic preferences, we expected that preference for default processing would be associated with higher answerability judgments and intellectual processing with lower answerability judgments. Although the earlier mentioned result showed that participants high in default processing and low in intellectual processing were tricked more by the illusion questions, only a weak correlation was found between intellectual processing and low answerability for the illusion questions when using the future scale. Although it was small, this correlation supports our hypothesis. Since people high in intellectual processing enjoy and take time to think about problems , it is llower answerability ratings on the non-consensus and the illusion questions for the current answerability scale. According to explanatory style or dispositional optimism. Explanatory style relates to how the individual tends to explain good or bad outcomes and may be less relevant to the present study. Dispositional optimism means a tendency to expect good things and few bad things to happen in the future, to believe that one can reach one’s goals  speaks against this interpretation, although this correlation, in spite of it being significant, was relatively low. It would seem more likely that the lack of associations between the consensus questions and the individual difference variables is a true effect in the sense that individual difference variables, for reasons discussed above, may have had less influence on the judgments of these questions, although future research should examine this issue further.As already noted, the results showed that answerability judgments of the consensus questions were not appreciably influenced by the individual difference variables. This could be due to a ceiling effect since the consensus questions received very high ratings, limiting the spread of their answerability ratings. However, the fact that the answerability ratings for the two types of questions were significantly correlated  can be argued to have a fairly positive attitude toward research and science. Future research should test if our results generalize to more representative groups of people and to people faced with uncertain but important questions, for example, researchers and politicians. Furthermore, future research should preferably use a larger number of answerability questions, especially a larger number of illusion questions.Finally, future research should also investigate people’s answerability judgments of other types of questions than in the present study. Examples are questions relating to current controversies in science and questions more clearly related to social life, such as questions whether a suspect can be considered guilty “beyond reasonable doubt” (see e.g., BK has with CMA had the main responsibility for data-collection, analysis, conclusions and creation of the manuscript. BK and SB have performed approximately half of the statistical analyses each. Furthermore, SB has actively supported the development of the study and has several times reviewed the paper and contributed to the Introduction and the Discussion with formulations and interpretations of theories.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.