This meta-analysis systematically evaluated the efficacy and safety of anti-PD-1/PD-L1 antibodies for pretreated advanced or metastatic nonsmall cell lung cancer (NSCLC) and investigated the correlation between PD-L1 expression levels and effectiveness of anti-PD-1/PD-L1 antibody. The methodology was based on the Preferred Reporting Items for Systematic Reviews and Meta Analyses (PRISMA) and the Cochrane Collaboration guidelines. Our research included five randomized-controlled trials involving 3,025 patients. We compared anti-PD-1/PD-L1 antibodies  with docetaxel in pretreated patients with advanced or metastatic NSCLC. The pooled hazard ratio (HR) for overall survival (OS) and progression-free survival (PFS) was 0.69  and 0.87 , respectively. Meanwhile, the pooled risk ratio (RR) for objective response rate (ORR) was 1.53  in all patients. Subgroup analyses showed that anti-PD-1/PD-L1 treatment significantly improved OS in patients with PD-L1 expression at any level, even in patients with PD-L1<1%. The RR for occurrence of grades 3 to 5 treatment-related adverse effects was 0.23 . OS, PFS, and ORR were significantly more improved for patients treated with anti-PD-1/PD-L1 antibodies than for those treated with docetaxel. Anti-PD-1/PD-L1 therapy resulted in longer OS than docetaxel, regardless of PD-L1 expression; however, higher PD-L1 levels were likely to correlate with better outcome. Anti-PD-1/PD-L1 antibodies also had a better safety profile than docetaxel. Lung cancer represents the main cause of cancer-related mortality worldwide, , 2 with Patients with previously treated, advanced, or metastatic NSCLC are difficult to treat, with systemic cytotoxic chemotherapy  having only modest benefits. In recent years, epidermal growth factor receptor (EGFR) inhibitor development and application has shown significant benefits for advanced or metastatic EGFR-positive NSCLC patients, –8 thoughImmunotherapy is a relatively new paradigm for the treatment of NSCLC. The programmed death-1 (PD-1) receptor, expressed by activated T-cells, is engaged by the tumor-expressed ligands PD-L1 and PD-L2 to reduce T-cell activation and facilitate tumor immune escape. –12 PD-1/Several inhibitors  targeting the PD-1 immune checkpoint pathway have been developed and approved by the United States Food and Drug Administration (USFDA) for the treatment of NSCLC. Compared with docetaxel, Nivolumab, a fully humanized IgG4 PD-1 inhibitor, showed significantly better overall survival (OS) and response rates (RR) in advanced squamous NSCLC, regardless of PD-L1 expression level . In anotIn the 2017 updates (Version 4), the NCCN Panel recommended that PD-L1 levels did not instruct the guidelines for treatment with some PD-1/PD-L1 agents, while other PD-1/PD-L1 agents were approved restrictively for patients with PD-L1 expression level ≥1%. Thus, the question remains whether PD-L1 expression should serve as predictor and guide for patient selection.The aim of this meta-analysis is to further evaluate the efficacy and safety of anti-PD-1/PD-L1 agents in advanced NSCLC patients. A subgroup analysis was performed to determine the correlation between PD-L1 expression level and clinical outcome and to establish guidelines for PD-L1 antibody treatment in patients with low or negative PD-L1 levels.This meta-analysis was performed in conformity with the PRISMA   and CochWe performed a literature search of PubMed, Embase, and Cochrane Library electronic databases, using a combination of the terms “Carcinoma, Nonsmall-Cell Lung” [MeSH] or “NSCLC” and “PD-1” or “PD-L1” and “nivolumab” or “pembrolizumab,” or “atezolizumab.” The last search was performed on March 20th, 2017. No restrictions for language or publication year were set in the search.The criteria for study inclusion were as follows:(1) Prospective randomized-controlled trials (RCTs) designed for PD-1/PD-L1 inhibitor therapy for patients with advanced or metastatic NSCLC that had been previously treated.(2) Published efficacy and safety measures reported and correlated to PD-L1 expression levels.Two reviewers (Q.L.Z. and X.H.W.) independently reviewed all abstracts, obtained full-text reports, and extracted data into separate databases. Disagreements were resolved through team discussion. For each study, the following information was extracted: first author's name, year of publication, trial phase, number of randomized patients, treatment strategies, clinical outcomes, PD-L1 status, hazard ratio (HR) for overall survival (OS) and progression-free survival (PFS) and their 95% confidence intervals (CIs), objective response rate (ORR), PD-L1 expression level, and overall grades 3-5 adverse events (AEs) and per grades 3-5 AEs.The primary endpoint was overall survival rate. Secondary endpoints included PFS, proportion of patients with an objective response rate (ORR), and safety. The analyzed safety outcomes were grade 3-5 adverse events (AEs), including fatigue, decreased appetite, nausea, vomiting, diarrhea, constipation, anemia, neutropenia, and febrile neutropenia.The 5-item Jadad scale was used to assess the quality of clinical trials and the calculated score was based on randomization, double-blinding, and reported withdrawals  20]..20]. P<0.05 was considered statistically significant. We assessed heterogeneity using a χ2 test with P<0.10 considered to be statistically significant. A fixed effect model was used when heterogeneity between studies was absent and a random effect model was used when heterogeneity was present. Subgroup analysis was calculated based on PD-L1 expression levels. Sensitivity analyses were used to estimate the effect of each individual study by removing one by one from analysis. Publication bias was assessed using funnel plots.All outcomes were pooled using RevMan 5.3 (Nordic Cochrane centre). Our analyses pooled HR with 95% CIs for OS and PFS and risk ratios (RR) with 95% CIs for ORR and grade 3-5AEs. HR<1 favored the experimental group (anti-PD-L/PD-L1 antibodies) whereas HR>1 favored the control (docetaxel). For each objective response rate and grade 3-5 AEs, a risk ratio (RR) was calculated based on the absolute numbers of patients presenting the objective response and grade 3-5 AEs, respectively. RR for ORR and AEs<1 indicated a higher overall response rate and toxicity in the control (docetaxel).A total of 101 relevant studies were electronically retrieved and 96 were excluded for the reasons shown in Five published RCTs involving 3,025 patients with subgroup analysis assessing the efficacy and safety of PD-1/PD-L1 inhibitors in NSCLC were included in this meta-analysis –17, 21. All included trials were considered high-quality data, as they were randomized when comparing anti-PD-1/PD-L1 agents  with docetaxel in the second or third line setting. Subgroup analyses, performed in all trials, explored the relationship between PD-L1 expression level and anti-PD-1/PD-L1 antibody efficacy. P<0.0001, and Ph=0.67)   (I2=59%), and the pooled RR for ORR was determined using a random effect model.Pooled results showed that anti-PD-1/PD-L1 antibodies significantly improved the OS   and PFS Ph=0.11)  in all pPh=0.03) . Moderat P=0.005, and Ph=0.29)  and ORR Subgroup analyses showed that anti-PD-1/PD-L1 antibodies could result in longer OS   than doc P<0.00001) ( P<0.00001), and ORR  when compared with docetaxel.However, in the PD-L1≥1% subgroup, PD-1/PD-L1 inhibitors significantly improved OS  , PFS   and for Therefore, this meta-analysis indicates that anti-PD-1/PD-L1 agents exhibited high efficacy in the treatment of advanced NSCLC. Anti-PD-1/PD-L1 therapy also had considerable activity for NSCLC and was superior to docetaxel in the PD-L1<1% population. PD-1/PD-L1 inhibitors tended to be associated with PD-L1 expression level. Higher PD-L1 expression was likely to be associated with increased benefit from anti-PD-1/PD-L1 agents.The meta-analysis showed that the rates of overall grade 3-5 adverse events (AEs) for the anti-PD-1/PD-L1 therapy were significantly lower than those of docetaxel . For anyThe direction and magnitude of the statistical significance of the overall results were confirmed by this analysis. The benefit of anti-PD-1/PD-L1 antibodies on overall survival was maintained (HR 0.69–0.69) even when we changed the fixed-effect model to a random-effect model. Subsequently, an influence analysis was performed by excluding individual studies. The benefit of anti-PD-1/PD-L1 antibodies on OS  did not vary regardless of study removal. The sensitivity analysis indicated the stability of all trials in the anti-PD-1/PD-L1 agents group.A funnel plot indicated no evidence of substantial publication bias .Blocking inhibitory immune checkpoints has recently gained interest as an immunological therapy for different kinds of cancer, especially advanced NSCLC. Binding of PD-1 to its ligands (PD-L1 and PD-L2), which are present on tumor cells, suppresses T-cell activation and results in immune response evasion , 22–24. In this study, a systematic meta-analysis of randomized clinical trials demonstrated the high efficacy and safety of anti-PD-1/PD-L1 antibodies for previously treated patients with advanced or metastatic NSCLC. Pooled results confirmed that anti-PD-1/PD-L1 agents significantly improved OS, PFS, and ORR in advanced or metastatic patients, both in the intention-to-treat population and in subgroups with PD-L1 expression level at 1% or more, 5% or more, and 10% or more. A high PD-L1 expression was likely to be associated with increased benefits. Furthermore PD-1/PD-L1 inhibitors also improved OS in the population with PD-L1<1%, which contradicts the guidelines for pembrolizumab administration only in PD-L1 positive patients (PD-L1≥1%). This discrepancy, may be attributable to the fact that the pembrolizumab trials excluded patients with PD-L1<1% patients or that the PD-L1 test might not accurately determine tumor PD-L1 levels. Other possible reasons may include heterogeneity of expression and sampling error, or that test samples predate earlier lines of therapy. Therefore, patients with PD-L1 expression levels just below and just above 1% will likely exhibit similar responses. Our results provide useful information for clinicians to inform their patients about treatment options for advanced NSCLC in the PD-L1<1% population. However, further research is needed to confirm these findings.Subgroup analyses showed a trend toward a greater efficacy as PD-L1 expression level increased. In other words, patients that expressed the highest levels of PD-L1 derived the greatest benefit from anti-PD-1/PD-L1 therapy . Importantly, patients with PD-L1<1% (HR=0.79) also experienced OS longer than those treated with docetaxel without any evidence of statistical heterogeneity; however, PFS and ORR showed no difference. These results may imply that the benefit from PD-1/PD-L1 inhibitor versus docetaxel in pretreated advanced NSCLC is not limited to the PD-L1>1% population. More importantly, our findings could indicate a dose-effect relationship between the levels of PD-L1 expression and the potential benefit from PD-1/PD-L1 inhibitors. However, our meta-analyses included only five RTCs, and to further confirm these hypotheses, larger sample size studies are necessary.I2=78%, Ph=0.0003). The reason for heterogeneity could be because different PD-1/PD-L1 inhibitors  have potentially specific pharmaceutical characteristics. Treatment-related AEs of grade 3-5 were similar with those observed after docetaxel treatment and included decreased appetite, nausea, vomiting, and constipation. However, hematological AEs (anemia and neutropenia), febrile neutropenia, fatigue, and diarrhea were all significantly less common for anti-PD-1/PD-L1 agents. Only a small percentage of patients treated with anti-PD-1/PD-L1 agents reported immune-related adverse events, including hypothyroidism and pneumonitis, and with the use of appropriate protocol guidelines, these events were relieved. Our study showed that anti-PD-1/PD-L1 therapy is superior to docetaxel in clinic application and presents a lower risk for treatment-related adverse events.Treatment-related adverse effects are an important evaluation index for any antitumor therapies. Our meta-analysis showed that anti-PD-1/PD-L1 antibodies had lower risk of total grade 3-5 adverse events than docetaxel. Pooled RR for total grade 3-5 adverse events was 0.29  compared with docetaxel, with statistical heterogeneity (In an era of personalized medicine using PD-1/PD-L1 inhibitors, predictors of response to therapy are important for making informed treatment decisions. PD-L1 expression may be an encouraging predictor for anti-PD-L1/PD-1 therapy in NSCLC, but standardizing PD-L1 testing has presented several problems. Different studies showed contradictory results regarding the relationship between drug efficacy and PD-L1 expression levels. Brahmer et al.  showed tOur analysis has some limitations. Due to the recent introduction of PD-L1 inhibitor therapy for pretreated advanced NSCLC, there were only five randomized clinical trials that investigated the efficacy and safety of anti-PD-1/PD-L1 antibodies, which limited the number of studies available for our meta-analyses. More randomized controlled trials with a larger sample size are needed to establish and replicate these clinical outcomes. Second, all five trials were liable to probable bias due to an open-label study design. Third, statistical heterogeneity was found when we pooled the objective response rate and the grade 3-5 adverse effect rates, respectively. Because of this, we used a random-effect model to pool outcomes. Since no evidence of substantial publication bias was found, these results indicated reliability to evaluate clinical outcomes.In conclusion, we analyzed five RCTs and systemically verified favorable OS, PFS, and ORR of anti-PD-1/PD-L1 therapy for pretreated advanced or metastatic NSCLC and demonstrated higher efficacy and safety for these agents than for docetaxel. More importantly, the results of this meta-analysis suggested that anti-PD-1/PD-L1 antibodies could also improve overall survival even when PD-L1<1%, which has not been recommended by previous studies. Our results could be of great value in guiding selection of clinical therapeutic regimens. More prospective studies are necessary to confirm these results and to improve the optimal dosage for PD-1/PD-L1 inhibitors in NSCLC.


In this research, we established a facile and low-cost strategy to construct graphene thin film enabled yarn sensors (GYS) by combining the process of graphene oxide (GO) coating and reducing on polyester (PE) wound spandex yarns. According to systematic processing-property relationship study, a key finding of this work discovers that the degree of resistance recovery as well as gauge sensitivity of GYS can be well controlled and modulated by a pre-stretch treatment. Specifically, as the level of pre-stretch increases from 0 to 60%, the deformable range of sensor that guarantees full resistance recovery prolongs evidently from 0% to ~50%. Meanwhile, the gauge factor of GYS is tunable in the range from 6.40 to 12.06. To understand the pre-stretch process dependent sensing performance, SEM analysis was assisted to evidence the growing size of micro-cracks determining dominantly the behavior of electron transport. Lastly, to take better advantage of GYS, a new wearing mode was demonstrated by direct winding the yarn sensor on varied portions of human body for monitoring different body movements and muscle contracting & relaxing. With continuous development, metallic strain gauges as well as other traditional sensing elements/interfaces/devices with mechanical rigidity and/or limited sensitivity are no longer the best candidate to be applied in smart devices11. In a sharp contrast, the next-generation sensors enabled by new materials and new nanotechnology are highly demanded to meet the rising standards of high sensitivity, mechanical flexibility, and wearable stretchability13. Toward this goal, carbon nanomaterials have been widely explored by virtue of their robustness, structural noninvasiveness, and multifunctional properties. Compared with other carbon family members, e.g. fullerene14, carbon blacks16, and carbon nanotubes18, graphene show multiple unique advantages, including the ease of aqueous colloids preparation, great conformability to substrates and interfaces, and advanced contact-breakage mechanism with excellent piezo-sensitivity, etc.21. The above-mentioned characteristics have guaranteed a variety of novel strategies for assembling varied types of graphene sensors, e.g., 1D CVD-grown nanofibers22, surface-coated fibers24, hybrid composite fibers25, 2D thin films and free-standing papers27, as well as 3D foams, sponges and aerogels28.Since the last decade, flexible and wearable electronics have been attracting considerable attentions with upgrading concepts and strategies in varied fields of applications, e.g. smart fabrics19. Following this line of thought, in combination with extraordinary properties of graphene, different 1D graphene fiber sensors have been proposed for wearable applications. For examples, Wang et al. applied CVD method to fabricate graphene/PVA core-sheath nanofibers with the gauge factor of 5.02 and stretchability up to 16.1%22. Using one-step wet spinning processes, Jalili et al. fabricated reduced graphene oxide fibers with ultimate elongation of ~3% for multifunctional textiles29. To meet larger stretching requirements for monitoring human-body movements such as elbow and knee joint actions30, graphene enabled composite fibers are demonstrating great advantages when using commercial elastic yarn as the core substrate. By coating thin graphene layers onto elastic fiber yarns through dip-coating and layer-by-layer processes, for instance, Cheng et al.31, and Park et al.32 respectively developed graphene covered PE/PU yarns and graphene/PVA coated yarns for monitoring a variety of human motions from head to foot. Montazerian et al.33 also applied a layer-by-layer process to coat graphene nanoplatelets on silicone sheathed spandex yarns with tunable sensing range >40%. Compared with CVD and wet spinning processed sensors, the composite fibers show better mechanical affinity to woven fabrics, superior tensile performance, and wider applications.When considering embeddable and wearable characteristics for flexible electronics, the sensor with unique one-dimensional format is expected to be more flexible, lightweight, robust, and straightforward to be assembled through winding, weaving, and/or printing processes34 to 56.335 with different assembling methods and deformation conditions. For another, it is lacking of a systematic work investigating the structure dependent sensitivity, deformability, and reliability, which is crucial for unveiling the sensing mechanism. With effective capability to alter and control different microscopic features such as micro-cracks36, wrinkles37, and packing densities35, the pre-stretch process has already become a commonly-applied strategy to tune the performance of varied nanocarbon sensors, including processing efficiency38, mechanical/electrical stability39, as well as sensing sensitivity37. Following this line of thought, the application of pre-stretch toward the understanding of process dependent sensing property and mechanism for the unique graphene enabled smart yarn is highly significant. However, the relevant study is still not appeared.As a new-emergent strategy for assembly of graphene 1D fibers, nevertheless, the sensing performance as well as the mechanism is far from being fully exploited, especially when the commercial yarn core is always specifically-strengthened by multiple sheathed fiber layers twisted/coiled through certain angles. With the overview of relevant works, for one thing, it is still not understandable about the variation of the reported gauge sensitivity varied noticeably from 1.5Motivated by the argument mentioned above and to further advance this emerging field, in this research, we systematically explored the unique pre-stretching treatment on regulating both electrical stability and gauge sensitivity toward understanding the sensing behaviors of graphene thin film enabled yarn sensors (GYS). Firstly, a facile and low-cost strategy was established to construct the graphene coating layer on polyester (PE) wound spandex yarns by combining the process of graphene oxide (GO) coating and reducing. Ascribed to the helically covered structure, it is rather interesting that the level of pre-stretch plays an important role on controlling the degree of resistance recovery. As the pre-stretch is increased from 0% to 60%, the ideal deformable range of the sensor that guarantees full resistance recovery prolongs evidently from 0% to ~50%. In addition to the enhanced working range, the pre-stretch dependent sensing performance is highly advisable to optimize the gauge sensitivity of GYS from 6.40 to 12.06. Based on microscopic explorations, the apparent processing-property relationship of GYS is further explained by the variable size of micro-cracks on graphene thin films bridging originally over PE loops, influencing dominantly on both axial and helical directions of electron transport. To take better advantage of lightweight, conformability, tunable sensitivity, and free from packaging medium, a new wearing mode has been demonstrated by direct winding GYS on varied portions of human body for monitoring different motions, such as finger bending and muscle contracting & relaxing.40, the Na2S2O4 solution is applicable by oxidizing S2O42− to SO32− through reacting with hydroxyl and epoxide groups in GO41. Once again, the effective reducing process was convincingly proved by the pitch-black appearance of GYS as shown in Fig. 3) as well as copper electrode connection, the as-produced GYS sensors were ready for the following structural analysis and performance evaluation.Figure 31. Under higher magnification, Fig. −1 and 1582 cm−1 for both GO and GYS, confirming the existence of graphitic structures. After the process of GO reduction, the increased intensity ratio of D and G peak (ID/IG) from 0.968 to 1.072 suggests both the decrease in average size of the sp2 domain as well as the increased quantity of structural defects42. In addition to Raman, XRD spectra show clearly the shift of the characteristic 2θ peak from 11.38° to 24.27° after the reduction process, reflecting the removal of oxygen-containing groups and the decrease of interlayer spacing from 0.778 nm to 0.360 nm which is fairly close to the one acquired from TEM (0.350 nm)43.In addition to visual inspection, structural and chemical details of GYS were further evaluated by varied microscopic and spectroscopic methods. Low-magnified scanning electron microscope (SEM) image Fig.  first un45. Through the measurement, sensitivity of the smart yarn can be quantified by determining the gauge factor (GF), which is defined as d(ΔR/R0)/dε where ε is the mechanical strain applied to the yarn, R0 and R are respectively the resistance of GYS before and after deformation. Since the length of the yarn cannot be completely restored when mechanical strain exceeds 60% ascribed to its viscoelastic behavior46, the upper limit of its working range is bound to this critical value for the ease of cyclic testing. Figure 0) accordingly varies as the enhancement of mechanical deformation from 0 to 5/20/40/60%, intriguing the piezoresistive behavior of the conducting layer. During the stage of releasing (dashed lines), nevertheless, the decreasing trend of ΔR/R0 is not able to follow its rising trace. More seriously, the resistance fails to recover back to its initial value regardless of the level of the subjected deformation. The inset picture also clears that the higher the degree of stretching, the greater is the resistance when totally relaxed with the zero-strain ΔR/R0 ranging from 2.6% (5% strain treated) to 11.5% (60% strain treated). To better quantify this behavior, a representative GYS specimen as produced by the spraying & reducing assembly was successively treated by different pre-stretching processes from low to high levels. As the pre-stretch level is steadily increased from 0 to 60%, Fig. In order to systematically evaluate the sensing property, coupled electrical - cyclic tensile tests were applied to acquire piezoresistive responses of GYS sensors0) after experiencing the corresponding treatment of pre-stretch. As shown in Fig. max − Rf)/(Rmax − R0) where Rmax, Rf, and R0 are respectively maximum, final, and initial resistance, Fig. Benefit from the cyclic test, it is worth noting that the pre-stretching process plays a dominant effect on recovery of initial resistance. It means that if a GYS sensor has experienced certain treatment of mechanical stretching, the irreversible issue of resistance could be weakened in next stretching cycles. To prove the idea, a comparison study was examined for investigating the behavior of strain dependent resistance change  by combining the process of graphene oxide (GO) coating and reducing on polyester (PE) wound spandex yarns. The effect of pre-stretch processes on sensing sensitivity and degree of resistance recovery of GYS was studied systematically. The research results show that proper pre-stretch can optimize the performance of GYS. The ideal deformable range of the sensor that guarantees full resistance recovery increases from 0% to 50% after applying a 60% pre-stretch. As for the sensing performance, the gauge sensitivity increases from 6.40 to 12.06 when applied by a 30% pre-stretch. The sensing mechanism was further explained by exploring the micromorphology. With the augmentation of pre-stretch levels, the size of micro-cracks on graphene thin film grows accordingly, introducing the change of electron transport behavior on graphene thin films. As contact resistance increases, GYS becomes more sensitive but shows negative effect when beyond a critical value, ascribing to the cut off of the axial transmission path of electrons. Based on fundamental study, a new wearing mode of the yarn sensor by direct winding GYS on varied portions of human body has been demonstrated for monitoring different motions, e.g. finger bending and muscle contracting. The unique characteristics of GYS sensors with lightweight, robust, tunable sensitivity, and free from packaging medium demonstrated in this work can be highly valuable for developing next-generation wearable electronics.The graphene oxide (GO) dispersion was prepared by sonicating a mixture of 600 mg of graphene oxide nanosheets which was synthesized via the Hummers method in 200 mL of deionized water in an ice bath using an ultrasonic processor (Ultrasonics FS-600 N). The ultrasonic processor was operated in a pulse operation mode  with the power set at 510 W for 3 h to obtain the 3 mg/mL GO dispersion.The spandex yarns wound around polyester (PE) are commercially available. The yarn with the cut length of 5.5 cm was cleaned repeatedly using deionized water and dried with a heat gun for 5 min. A pair of clamps was used to fix the yarn sitting on the platform of the spray-coating system, consisting of a heat gun, an air compressor (Iwata IS-50), and an air brush (Iwata HP-CS) equipped on a computer controlled moving stage (Nordson E3). The heat gun was used to accelerate solvent evaporation and the air compressor was used to provide the pressure needed for spraying. In view of the requirement to avoid the heat gun wind for disrupting the spraying process, the heating machine was placed perpendicular to the air brush. To control the heating temperature of the sample around 100 °C, the distance from the heat gun to the yarn is set at 18.5 cm. Additionally, the distance from the nozzle to the elastic substrate is about 6 cm for best spraying effect. Before spraying, the yarn was slightly stretched to prevent it from shaking violently during the coating process.With the back-and-forth nozzle moving speed of 2 cm/s and spraying rate of 2.5 mL/min, 5 mL as-prepared GO dispersion was sprayed on the yarn surface with the accompanied color change from white to brown. After the top-direction spraying was completed, the yarn was axially rotated by 180° and received another 5 mL spraying treatment to ensure the uniform coating of graphene oxide. Finally, the GO coated yarn was soaked in 90 °C pre-heated 0.06 mmol/L sodium hydrosulfite solution for 2 h and 30 min. The reduced GO (rGO) yarn was washed with deionized water for several times and dried before use.The SEM characterization was accomplished using a JEOL JSM-7001F FE-SEM. Raman spectra were recorded on a HORIBA LabRAM HR Evolution spectrometer. XRD characterization was performed on Rigaku D/max 2550 with Cu Kα radiation (λ = 1.54°). In addition, the yarn was sputter coated with platinum before being examined by FE-FEM.To facilitate measurement and reduce contact resistance, two copper pieces were pasted onto each end of the yarn using conductive silver paste before strain sensing tests. The electric properties of the smart yarns were measured by a Keithley 2510 source meter. The sensing performances were evaluated by a mechanical-electric coupling test including a universal testing machine (MTS E44.104 with 500 N load cell) and the source meter. The tension was controlled by the universal testing machine with the fixed deformation rate of 20 mm/min and real-time changes of the sensor resistance were recorded by the source meter programmed by LabVIEW. The sensing performance under other deformation rates was summarized in Supplementary Fig. Video S1Supplementary Document


Nanoscrolls (NSs) assembled from two-dimensional nanosheets have emerged as a novel type of one-dimensional nanomaterials because of their unique topological features and properties. The scale-up preparation of the NSs is crucial for their foundational and applied research. Herein, we report a general and straightforward approach for efficiently converting two-dimensional nanosheets into the NSs with high yield. We demonstrated the converting process by illustrating the formation of the graphene nanoscrolls through characterizing their morphology and structure using a scanning electron microscope, transmission electron microscope, Raman spectra, and X-ray diffraction spectra. The graphene sheets with a few-lay number were converted immediately and entirely into the graphene nanoscrolls when they mixed with an ethanol solution of silver nitrate at room temperature. The as-prepared graphene nanoscrolls were confirmed to be formed via the layer-by-layer assembly of graphene triggered by silver cyanide formed in site. Also, we extended this approach to construct the nanoscrolls of the hexagonal boron nitride, molybdenum disulfide, and tungsten disulfide, respectively, from their corresponding two-dimensional nanomaterials. In a broader context, this approach paves a significant new way for the large production of the NSs with cost-efficiency. Since the discovery of graphene, the carbon nanoscrolls was convinced of deriving from scrolling of graphene, named as graphene nanoscrolls (denoted GNSs)4. GNSs inherit not only excellent properties of graphene, such as high mechanical strength, outstanding electrical conductivity, and superior carrier mobility but also possess open topological structure at both ends and interlayer galleries5. It endows the GNSs with wonderful features, such as excellent electronic and electromechanical properties7. Thus, the GNSs have significant potential applications in energy storages, photovoltaic cells and sensors10. Besides graphene, recently it was reported that the other layered two-dimensional nanomaterials (denoted 2DNMs) could be rolled into the nanoscrolls (denoted NSs) too, such as h-boron nitride (h-BN), tungsten disulfide (WS2), and molybdenum disulfide (MoS2)11. The NSs have become a new type of one-dimension nanomaterials with novel properties. Therefore, the preparation of the NSs has attracted increasing attention of scientific researchers. However, most of the research work is limited to theoretical prediction and calculation15. Only a few previously published articles involved fabricating the NSs. Moreover, the reported processes are complicated, and the conversion yield is tiny limiting their application23. Up to date, there is no general and efficient method to enable assembling the NSs with high-yield.Carbon nanoscrolls were discovered when making multiwalled carbon nanotubes2, and WS2 by verifying the morphology and structure of the formed NSs using scanning electron microscope, transmission electron microscope, Raman spectra, and X-ray diffraction spectra. Also, the mechanism of the self-assembly scrolling processes is elucidated.Herein, we report a facile and general approach for production of high-quality NSs with high-yield from two-dimensional nanosheets. The NSs are fabricated via self-scrolling of the 2DNMs in the solution of silver nitrate and ethanol. We have demonstrated the formation of the nanoscrolls of graphene, h-BN, MoS3 ethanol solution when they were mixed and magnetically stirred. After stirring the solution for some time, the grey dispersion gradually changed into a clear solution  and a transmission electron microscope (TEM) were applied to characterize the microstructure and morphology of the GNSs. The SEM image Fig.  indicate−1) and the shift of the 2D peak are often used to judge the layer number22. Accordingly, the graphene sheets which scrolled to the GNSs are a few layer numbers which are agreement with the results characterized by TEM and AFM shown in Fig. 25.Raman spectrum is an excellent way to characterize the structure of the nanostructured materials. Comparing the Raman spectrum of the graphite, the graphene, and the GNSs shown in Fig. In order to explore the conversion process of the graphene to the GNSs, we have examined the change of the morphology and structure of the graphene with the reaction period via the SEM characterization. Figure 2, and WS2 to explore its generality. The SEM images , AgNO3 were purchased from Sinopharm Chemical Reagent Co., Ltd (China). Carbon dioxide (99.9%) was obtained from Shanghai high-tech Co., Ltd (China).Graphite powder was purchased from Sinopharm Chemical Reagent Co., Ltd (China). H-BN bulk powder was obtained from Strem Chemicals. The MoS2, and WS2 were exfoliated using a shear mixer in a supercritical CO2 approach similar to the previously reported paper26. The 1 g raw material was put into the reactor, and carbon dioxide was pumped into the reactor by the manual pump. When the pressure and the temperature reached a preset value of 12 MPa and 45 °C, the shear mixer was started and remained at a speed of 3000 r/min for 1 hour. The resultant sample was dispersed by an ultrasonic water bath and centrifuged. More than 80% of the exfoliated 2DNMs were confirmed to be 1 to 5 layers by AFM, TEM and Raman spectra.Graphene, h-BN, MoS3 was dispersed in ethanol solvent in a predefined mass ratio. When the dispersion was magnetically stirred for 30 min at room temperature, the 2DNMs rolled up and deposited as shown in Figs A certain amount of the exfoliated 2DNMs and AgNOScanning electrode microscope imaging was performed using an FEI Nova NanoSEM 450 (USA). Transmission electron microscope images were obtained using JEOL JEM-2100 TEM (Japan) at an acceleration voltage of 120 kV. The X-ray diffraction (XRD) recorded on a BRUKER D8 Advance X-Ray Diffractometer (Germany) using a Cu Kα radiation source (λ = 1.5418 Å). The Raman spectroscopy was recorded on a Raman Microscope excitation (USA) with a wavelength of 532 nm. Zeta Potential was performed with a Malvern Instruments ZS90 Particle Size and Zeta Potential Analyzer (UK).Supplementary Information


In the course of this developmental process, some L2 skills may eventually match or even overtake the corresponding skill in the L1. This shift in the balance between L1 and L2 is the focus of investigations of language dominance. The present study explores language dominance in four migrant populations . Investigating both the development of formal/controlled skills and more automatic aspects of lexical access and fluency, we aim to attain an understanding of how extralinguistic factors contribute to the development of both languages. We argue that an integrated perspective can contribute more profound insights into the predictors of this complex process of bilingual development. In particular, our findings show that statistical models based on linear relationships fall short of capturing the full picture. We propose an alternative method of analysing data, namely discriminant function analysis, based on a categorisation of the populations, and demonstrate how this can enhance our understanding. Our findings suggest that different aspects of the bilingual experience contribute differently to language development, regardless of language combination and type of skill measured. Contrary to what previous research suggests, measures relating to the intensity of informal use of both the L1 and the L2 in daily life are important in determining whether someone is a good or a poor L1 maintainer, while high vs. low success in acquisition appears to be predominantly associated with personal factors such as educational level.Late bilinguals who spend (part of) their adult lives in an environment where a language other than the one they learned in childhood is spoken typically experience a range of language development phenomena. Most obviously, they will acquire some level of receptive and/or productive knowledge of the new, or second, language (L2). How basic or advanced that level will be is determined by a range of environmental, experiential, attitudinal and individual factors. Secondly, they will most likely find the knowledge of their native language (L1) beginning to diverge from that of monolingual speakers in their country of origin, a process known as  Language dominance is an extremely complex concept, encompassing a wide range of aspects and features e.g., . These fx-axis representing language X (Lx) and a y-axis, representing language Y (Ly) 1. A speaker who is mapped close to the diagonal of this space  is someone whose proficiency is more or less ‘balanced’ between the two languages, while one who is closer to one axis than the other is ‘dominant’ in the ‘stronger’ language, the one in which s/he has scored more highly. It should be noted that this visualisation is a simplified and idealised one: ‘proficiency’ cannot easily be reduced to a single measure , and the position of the same individual may therefore vary considerably depending on what skill or task is being measured . Time, in this model, is the only dimension which is linear and unidirectional. Linguistic development is neither: it can shift toward the higher or the lower end of the spectrum in either language, encompassing both acquisition and loss. Such shifts can occur in bursts or slowly and gradually, they can reverse direction from growth to decline and back again and they can affect mainly one language, both languages equally, or both languages orthogonally .The third dimension in the imaginary space defined by proficiency in both linguistic dimensions in order for such models to fully capture the phenomenon and not to reduce the analysis to the development of one language only, nor to collapse them into a one-dimensional function, e.g., by subtracting one from the other.The formidable task for investigations of language dominance, then, is to provide explanatory models capable of mapping out how the predictor variables may interact and determine the intensity and direction of developmental changes for any given aspect of language proficiency over time. Crucially, we argue here that it is important to take into account Ly begins while Lx is still at an early stage of development. Findings indicate that the highly active phases of language development during childhood and adolescence allow great flexibility when it comes to shifts in language proficiency and dominance, with changes in external circumstances or exposure – such as the start of (nursery) school, a move between countries, or a return to the home country – often causing spurts of development and/or regression that may fundamentally change the overall multilingual balance within months  which argues that the addition of a new language will inevitably lead to changes in all of those aspects that we subsume here under the term proficiency in the language that is already established. A growing body of research provides evidence of such attrition in immersed as well as non-immersed bilinguals  occurring after the development of the L1 has reached a relatively mature level and L1 development has hence slowed down considerably. That being the case, the majority of investigations of late bilingual development focus on the L2, the assumption being that a level of stability – some kind of ‘steady state’ – has been reached in the L1 which makes it uninteresting for research e.g., . This nols e.g.,  and attels e.g., .X will grow at the expense or to the detriment of Language Y. Research on bilingual development has long since demonstrated that there is no such straightforward or trivial relationship  a loss in the other. When it comes to simultaneous or early bilingualism, popular understanding often has it that Language ip e.g., . BilinguWhen it comes to L1At, findings on the relationship between proficiency in the speaker’s two languages are quite sketchy. In early research a relatively straightforward correlation was often assumed, according to which “[t]he greater the degree to which a speaker masters one system, the greater the extent to which one might expect it to affect another system,” so that “we expect L1 loss to be greatest in individuals who […] have mastered the L2 to a relatively high degree” , p. 191.There are few studies to date which investigate proficiency in both the L1 and the L2 of late immersed bilinguals. The findings that do attempt to make direct comparisons across languages and tasks e.g.,  or link Figure 1 above into three sections  uses four quadrants: dominant bilinguals (either Lx < Ly or Lx > Ly), low-achieving balanced bilinguals and high-achieving balanced bilinguals. input and output are among the most important predictors  while for others it is more difficult to see how crosslinguistic equivalence can be established  are therefore problematic. Instead, the outcome variables should be selected to represent relatively general and holistic aspects of language proficiency, taking into account the extent to which these may vary crosslinguistically in both native and non-native populations.Studies of language dominance, which attempt to assess relative levels of proficiency across languages, first of all have to acknowledge that some of the skills or measures listed above lend themselves more readily to direct crosslinguistic comparisons of Given that L1At populations command two languages which are learned under similar conditions  but at different stages in life – that is, speakers who unambiguously have one native and one non-native language – a fruitful framework for the assessment of proficiency is the model proposed by (a)in combination with (c) the automaticity with which these types of knowledge can be processed  the largely explicit, conscious knowledge in the lexical domain (form-meaning mappings), and is restricted to those lexical items and morphosyntactic structures in spoken language which all adult L1 speakers share  and which they may encounter in all communicative situations. HLC, on the other hand, refers to more complex domains of use, encompassing less frequent items and structures as well as written language, and it is in this domain that native as well as non-native speakers vary considerably from each other. ‘Frequency,’ in this framework, is operationalised on the basis of the assumption that lexical items and grammatical structures follow a Zipfian distribution in naturally occurring language, where ‘highly frequent’ items (those belonging to BLC) are situated on the steep left side of the slope, while less frequent items are to be found on the flattening part of the curve to the right :22ff.Among the fundamental assumptions of the model are(a)Command of BLC should be homogenous within as well as across native populations, while these populations will be stratified when it comes to HLC.(b)Non-native populations will exhibit considerable variability with respect to both; they are unlikely to fully acquire BLC in their L2 beyond the domains of vocabulary but may nonetheless reach high native levels of HLC(c)Development of BLC in non-natives and HLC in both populations will be modulated by a combination of external and background factors :52f.2 who will exhibit variability in both BLC and HLC. Attriting populations, however, are similar to late L2ers in that they diverge from monolingual control populations in both domains in their L1, indicating that even for native speakers, becoming bilingual will affect performance on skills belonging to both BLC and HLC.Hulstijn’s model thus assumes two types of speakers: the ‘native speaker,’ who will be at ceiling for all components of BLC but may vary with respect to HLC, and the ‘non-native speaker’3:There are a host of findings demonstrating that many of the language components which belong to BLC, and which are therefore assumed to show little variance among ‘native speakers,’ are subject to change and L1 attrition in immersed late bilinguals. These includeAccentedness: while monolingual populations are typically perceived to be at ceiling in global foreign accent rating experiments, several studies have established an increase in variance of such ratings in immersed late bilinguals which can lead to some attriters being perceived as unambiguously non-native e.g., : ch. 4 aFluency: in both free and elicited discourse, L1At populations have consistently been demonstrated to be less fluent than monolinguals, as indicated by a slower speech rate and higher incidence of pauses, filled pauses, repetitions and self-corrections e.g., .Lexical access: L1At populations are less productive at generating lexical items in Verbal Fluency tasks e.g.,  and slowOvert/null pronouns: This is an example of grammatical features which form part of BLC. There are a large number of investigations demonstrating that overt pronouns come to be overgeneralised to contexts where monolingual natives would use null pronouns by attriters of pro-drop languages such as Bulgarian, Italian, Greek, and Spanish e.g., .C-tests or cloze tests, e.g., Given that even monolingual native populations are assumed to be stratified with respect to HLC, it is hardly surprising that this variance increases under the cognitive demands of bilingualism. A wide range of studies have demonstrated this, for example, for complex and infrequent syntactic phenomena. For example, several studies of embedding structures in L1 Turkish have found attriters to diverge most from monolinguals on those types of embedding which are morphologically the most complex  and occur least frequently in free speech e.g., . In a siIn addition to distinguishing between BLC and HLC, Hulstijn’s model furthermore differentiates ‘core’ and ‘periphery’ aspects of language proficiency: core components refer largely to linguistic knowledge and the speed with which it can be processed, while peripheral components refer to the more metalinguistic skills, such as interactional ability and knowledge of the characteristics of different types of (spoken or written) discourse , p. 41. The findings presented above suggest that the BLC/HLC model can profitably be extended to include the development of language proficiency under conditions of L1At. The fact that the model assumes a global and holistic approach to defining language proficiency furthermore makes it ideally suited for an investigation of the relative development of proficiency in both the L1 and the L2 within the framework of language dominance, as the components of the model can be assessed and compared across languages. The findings presented above illustrate that, for linguistic features in both domains, attriting populations develop increased variability and diverge from the native baseline.What remains entirely unclear, however, are the conditions or predictors which drive these changes: In all of the studies listed above, some of the attriters remain within the native range while others fall squarely outside it, but assessments of the impact of predictor variables have remained largely inconclusive e.g., . In otheThe present study sets out to test the hypothesis that the explanatory potential of investigations of language dominance can be enhanced by adopting a fully two-dimensional approach which takes into account performance in both L1 and L2. We furthermore assume that linear statistical models – that is, models based on regression slopes – may not be able to capture the complex interaction of different features of proficiency, personal background, exposure/use, and attitudes , and propose that a classification into different types of language developers may allow a more detailed picture to emerge.We ask the following research questions:RQ1:What is the predictive power of external factors for measures of language proficiency in both the L1 and the L2 of long-term immersed bilingual populations in a multifactorial linear model?RQ2:To what extent can the predictive power of external factors for measures of language proficiency in both the L1 and the L2 of long-term immersed bilingual populations be increased by adopting a data categorisation approach which does not rely on the linearity of relations?RQ3:What are the differences between HLC and BLC aspects of proficiency with respect to RQs 1 and 2?The data reported here were collected in 2004 (Study 1) when the PI (the first author of this paper) was affiliated with the Vrije Universiteit Amsterdam and in 2007 (Study 2) when both authors were affiliated with the University of Groningen. At this time, the humanities faculties at these institutions did not have a protocol for ethical approval nor an ethics committee, and there were no national guidelines in relation to this. All participants did provide written informed consent prior to the experiment. With hindsight we recognise the lack of formal ethical approval for the studies to be a shortcoming – which, unfortunately, cannot be addressed retrospectively. However, all of the materials and experiments reported on here have been used by both authors in subsequent investigations for which ethical approval was duly granted according to the protocols required by different institutions, including the University of Groningen, the University of Essex and the Humboldt University, Berlin. We are therefore convinced that the research design in itself is unproblematic from an ethical point of view.n = 106) with between 9 and 58 years of residence (LoR) in the Netherlands  and the Greater Vancouver area, Canada . Study 2, investigating aspects of Basic Language Cognition (BLC), was conducted with 87 migrants with between 10 and 43 years of residence in the Netherlands. 52 of these speakers were Turkish natives (mean LoR 22.57), while 35 were native speakers of Moroccan Arabic (mean LoR 23.31).The data for the present study were collected from four experimental and three control populations. Study 1, which focuses on the development of Higher Language Cognition (HLC), was conducted with native German speakers  collected similar data in the L2. In order to induce a predominantly monolingual language mode for these experiments, we considered it important that the researcher should be a speaker of the language which was the focus of the experimental session  with no knowledge of the other language. The researchers conducting Session 1 were recent arrivals to the Netherlands and native speakers of Turkish and Moroccan Arabic, respectively. Session 2 was conducted by two research assistants who were native speakers of Dutch but had no knowledge of either Turkish or any variety of Arabic. Unfortunately but inevitably this led to some participant loss, with data from only 63 participants available at Session 2. In this and the following sections, the dataset comprising 87 participants will be referred to as ‘Full dataset,’ while the dataset comprising the 63 participants with L2 data available will be referred to as ‘Limited dataset.’ 4. The questionnaire comprises a total of 77 questions in different formats: open questions , Likert-scale questions , and interval questions . The questionnaire and its coding and analysis are described in detail in Supplementary Table S1.Data on participants’ biography, language learning history, language use and language attitudes were collected by means of the same questionnaire in both studiesAll participants were asked to rate their proficiency in both their L1 and L2 (used here to refer to the language of the country in which they were living at the time of data collection) both at the time of migration and at the time of the interview, and also to state which of these languages they felt was the stronger at the present time. There were several interesting differences between groups, such that regardless of L1 background almost none of the migrants to the Netherlands knew more than a few words of Dutch before arrival while more than half of the migrants to Canada rated themselves as intermediate or proficient in English at arrival. At the time of testing, most of the bilinguals felt they had intermediate or good proficiency in the L2, with the English L2 speakers again standing out. The Germans rate their proficiency in Dutch more highly than the Turks and the Moroccans, possibly reflecting the advantage the close typological relatedness between their L1 and their L2 gives them. With very few exceptions, everyone rated their L1 proficiency at migration as ‘good’ or ‘very good,’ but that proportion dropped across the board for proficiency at the time of testing, although only one single speaker described it as ‘bad.’ Only among the L2 English speakers did more than a quarter of participants feel that their L2 had become stronger than their L1, while just over half of all L1 Germans thought both languages were equally good. Balanced bilingualism or dominance reversal were much rarer among the Turks and the Moroccans, with strong majorities in both groups feeling that their L1 remained their stronger language.The questionnaire contains a total of 25 5-point Likert Scale questions on frequency of L1 exposure and use:• L1 and L2 use within the family • L1 and L2 use with friends and acquaintances (4 questions)• L1 and L2 use at work (2 questions)• Frequency of use of L1 media • Frequency of visits to the home country (1 question)• Overall contact with and use of the L1 (2 questions)Some of these items had to be excluded for the present analysis as variability was too low . The general picture which emerged across these questions was that most participants continued to use their L1 on a fairly regular basis, the Germans slightly less so than the Turks/Moroccans. All groups had good social contacts within their new country but there was some variance, with roughly two-thirds of Turks and Moroccans reporting more friends who shared their L1 while over half of the Germans said their social network was composed mainly of native speakers of the L2. When it comes to language use in the family, the Turks stand out somewhat from the other groups with a much stronger claimed adherence to an L1-only policy, while half of the Germans report using L2 only . The Moroccans appear to occupy an intermediate position. A similar picture appears across most language exposure and use questions: most participants appear to have a fairly clear preference for one language in each context, with the Turks and Moroccans leaning more toward the L1 than the Germans. The only exception to this is the use of the native language for professional purposes, which stands at around 20% for both German groups but is quite rare for the Moroccans and Turks.Where attitudes toward the native language are concerned, the views seem more homogenous across groups, with over 75% in all groups saying it is important or very important to them to maintain their L1 and almost the same proportion of respondents saying it is important to them to pass it on to their children. An interesting finding emerged from the question “Which language do you prefer?”: while the Germans in Canada were split roughly evenly between their L1 and their L2, and three quarters of the Turks and Moroccans stated a preference for their L1, the only group that had a substantial proportion of self-reported balanced bilinguals for this questions  were the Germans in the Netherlands. This suggests that the similarity between L1 and L2 for the German–Dutch bilinguals may have facilitated the perception of a more balanced bilingualism. Interestingly, while there was only one speaker who reported ‘no preference’ on this question in the German–Canadian group, over 50% responded in the affirmative when asked whether they felt they were balanced bilinguals, while for all other groups, the answers across both questions seemed to be largely consistent.The overview of findings presented here points to two general problems concerning personal background data in language attrition research. Firstly, there are many questions with potentially important information for which there is missing data from a substantial proportion of informants – for example, not all participants have a partner and/or children. Secondly, as pointed out above , the data are not normally distributed: for most of the variables reported here, there is either a skewness toward the L1 end of the scale or a bimodal distribution. Both phenomena are a natural and inevitable characteristic of attrited populations: most studies find sustained preference for the native language, particularly where it comes to self-assessed proficiency. Furthermore, as is predicted by the Complementarity Principle , 2016, mIn order to alleviate these problems as well as reduce the number of predictor variables for analysis, we conducted a Principal Component Analysis (PCA). We addressed the problem of missing values by replacing them in each case with the neutral point on the scale. We chose this strategy over the more common approach of imputing missing values based on the rationale that, for the data in question, values either above or below the neutral measure would (incorrectly) suggest that the relevant language  played a role in the prediction of the outcome variables while setting it to neutral allowed the case to be included in the analyses without such an effect.age at testing) set to 1 and the minimum value set to 0. The PCA  identified a total of six components which were saved as factors (see Supplementary Table S2 for the full component matrix). The first component comprised 9 factors relating largely to the frequency of casual and informal use of the L1 and the L2, that is, with family and friends (Cronbach’s α = 0.881) and was labelled Interactive Use. The second component, Personal Background, comprised the variables age, length of residence and education (α = 0.609). The third component related to Perception and comprised the answers to the questions about current L1 proficiency and whether that had changed since immigration (α = 0.455). Component 4 comprised the Attitude-related variables of importance to maintain the L1, transmit it to the children, and culture of preference, alongside the frequency of use of L1 media  (α = 0.627). Overall Contact with the L1 was a unifactorial component, while ProfessionalUse of both L1 and L2 made up the last component (α = 0.476).All variables were standardised to the same scale prior to entry into the PCA, with the maximum value in the dataset  = 0.091, p = 0.001] and a more pronounced positive one for ProfessionalUse . These variables were log-transformed after a constant was added to make all values positive and the scale for the negatively skewed component was inverted. The transformed components were no longer skewed .The first four components were normally distributed. There was a slight negative skew for The data collection for both studies included a native language control group for the L1 tasks , and Study 2 also used a Dutch native control group for the L2 tasks. Controls were matched with the relevant experimental populations for age, gender, educational background and, in Study 2, region of origin within the L1 country. It is not the purpose of the present investigation to probe into issues of general proficiency or overall attainment against an idealised monolingual baseline, but to assess to what degree development has taken place in both languages within the proficiency space defined by the performance of the immersed bilingual population. While the descriptive statistics given below include the results from the control group as indicative values, they were therefore not used in the inferential statistics.5: a C-Test and a detailed self-assessment, each in both their L1 and their L2. Each of the two C-Tests comprised five short texts with a total number of 100 gaps determined by the schema proposed by In Study 1, participants completed four tasksC-Test in the second language was different for the two populations  the results were standardised for both groups by setting the lowest score in either population to 0 and the highest to 100. None of the tasks differed significantly across populations, although the Can-Do scales for the L1 approached significance, with the Dutch L2 speakers rating themselves somewhat higher than the English L1 speakers. The results are summarised in Table 2.Since the p < 0.01 for all four variables, and visual inspection revealed all of them to be negatively skewed. The variables were therefore inverted and log-transformed, which resolved the normality issue for all except the L2 C-Test. That variable was root-transformed instead, resulting in a normal distribution. All variables were subsequently re-calculated to the same scale, so that in all cases the lowest score achieved was set to 0 and the highest to 1. The resulting standardised scores all correlated with the original scores above 0.95 . Lastly, we calculated an average score for both tasks in the L1 as well as in the L2 .Tests of normality  were significant at Table 3).Study 2 also used four instruments: Firstly, there were two Picture Naming Tasks  in which participants were asked to say aloud the name of 78 objects which they saw as line drawings on a computer screen. The pictures were selected from the In this case, the L1 naming latencies of the two bilingual groups did not differ substantially from each other. Nevertheless, in order to ensure that language-specific differences would not impact on the results, we followed the same strategy for standardisation within the language groups as described above under Study 1 (based on the Full Dataset). L2 naming latencies were standardised only on the basis of the bilingual data and did not include the monolingual data.The second set of variables was derived from a semi-structured interview conducted by a native speaker of the language in question  with no knowledge of the other language. Both interviews were autobiographical, informal, and focused on different aspects of the emigration experience. All interviews were transcribed and coded according to the guidelines set out in the CHILDES project for the CHAT format . The fol• VOCD . For the• Fluency: for each participant, the number of filled pauses, repetitions and retractions (self-corrections) was counted and subsequently standardised to incidence per 1,000 tokens. Fluency variables were standardised across the groups for all speakers in the same way as described for the other variables above and averaged to one overall fluency measure per speaker in L1 and L2.Tests of normality (K–S) showed no deviations from the normality assumption for any of the above variables. All were standardised to the same scale and direction (0 being the worst attained score and 1 being the best), and subsequently, one average measure for L1 and L2, respectively, was created by averaging the three subtasks.In order to assess to what extent the variables established above for personal background, use of L1 and L2, and attitudes may be used to predict the outcome variables we conducted two analyses for each dataset: a multivariate analysis of covariance (MANCOVA), which creates an overall model but also allows identifying the regression slopes associated with each predictor for each outcome variable (RQ1), and a discriminant function analysis (DA), which allows a non-linear assessment of the impact of the predictors (RQ2). Each analysis was conducted separately for HLC aspects of proficiency (Study 1) and BLC aspects of proficiency (Study 2) (RQ3).C-Test L1, C-Test L2, Can-Do L1 and Can-Do L2. The six components identified by the PCA were entered as covariates. All components were entered together. Roy’s Largest Root was significant for all components except Contact (see Table 4 for the full results). Interactive Use was significantly associated with both of the L2 measures, with a higher level of L1 use associated with a lower L2 C-Test score and L2 self-rating, but did not influence outcomes in the L1. The Personal Background component was associated with both self-assessments, a higher score on this component  leading to participants rating themselves better in L1 and poorer in L2 – without, however, this being reflected in the more objective proficiency scores yielded by the C-Test. Perception influenced both L1 measures, with participants who claimed their L1 had not changed and was at a high level at the present time achieving better scores on the C-Test and rating their proficiency more positively. Attitude had the most consistent effect: participants for whom the maintenance of their L1 and its transmission to the next generation was more important and who stated a preference for their home culture achieved better scores on the L1 C-Test and also rated themselves more positively in both languages. A higher level of use of the L1 for professional purposes and a lower level of use of the L2 was associated with higher C-Test scores in both languages. For the full models, effect sizes were medium to large, partial η2 values ranging from 0.22 to 0.33, while the individual associations were weak at best with partial η2s between 0.05 and 0.2.The first MANCOVA was conducted on the four standardised outcome variables from Study 1: Interactive Use and Personal Background variables only became significant for the L2, while Perception was significant only for the L1. Attitude and Professional Use were significant predictors in both models, while Contact was not significant for either language. Except for the impact of professional L1 use on the L1, effect sizes were considerably lower than in the full model, ranging from 0.06 to 0.22 (see Supplementary Table S3).In order to assess to what extent including both languages in the model improves explanatory validity (RQ2), we repeated the MANCOVA twice. The first model included only the L1 measures as dependent variables, and the second only the L2 measures. Here, Interactive Use, with a higher level of L1 use in informal contexts associated with slightly slower responses on the L1 PNT as well as a reduction in L1 disfluencies. Partial η2s were around 0.11 (weak effect) for the individual measures and 0.27 (medium effect) for the overall model. A few other significant relationships were observed (see Table 5), but in those cases the overall model was not significant. These findings confirm earlier studies showing that accounting for informal features of language attrition on the basis of background variables is highly problematic in models based on linear regression slopes .Repeating the MANCOVA for each language separately did not return a significant result for any of the predictors except The findings from these two analyses are interesting in the light of previous investigations of L1 attrition in that they underscore that, while including results relating to performance in both languages can increase explanatory adequacy, analyses looking for linear regression slopes typically yield few results and have little predictive power. While the analyses of the HLC skills presented above are overall significant, the predictive power of the independent variables is limited and inconsistent, and effect sizes are weak. The situation is even worse with respect to BLC aspects of proficiency, where no coherent picture emerges at all.linear trends and correspondences in the data . In other words, any one predictor will only be revealed as significant if its impact on the outcome variable is the same for all or most of the participants. This can be seen, for example, in the fact that Professional L1 Use has the strongest impact on formal tasks such as the C-Test: it makes sense that individuals who engage with language as part of their job would develop enhanced awareness of style, orthography etc., facilitating these kinds of tasks. However, this relationship may not hold for all speakers  and may be far more complex for other types of background variables. For example, it is possible that some factors may interact with each other in non-linear ways , indicating that participants who perform well in one language tend to perform well in the other , but there are quite a few marked exceptions to this trend, as visualised in Figure 2A: for example, the two crosses toward the bottom right of the panel represent two participants who are among the lowest performers in the L1 but score within the top 15% in the L2. For the less controlled aspects of language use belonging to BLC aspects of proficiency, such as lexical access measured by the PNT and VOCD, on the other hand, no significant correlation between languages obtains, and no overall pattern can be detected from the scatterplot .A more serious problem for linear analyses is the fact that, as pointed out above (see section “Investigations of Development in Both Languages”) the balance between the L1 and the L2 is not the same for all members of the population: some speakers may preserve excellent proficiency in the L1 while also excelling in the L2, while for others, the development or maintenance of one language may come at the expense of the other, and others still may regress in their L1 without ever reaching very advanced levels of the L2. This variance cannot be captured by single-dimensional approaches, where the score obtained in one language is subtracted from that of the other, but it also eludes analysis by linear modelling. Consider the scores obtained on the median split of all participants in Study 1 and in Study 2. This was calculated on the basis of a single average score per language for the standardised variables C-Test and Can-Do Scales in Study 1 and PNT, VOCD and fluency in Study 2. Each participant was then categorised as having scored either above or below the median in each language, yielding four groups (see Table 6). The division is visualised in Figure 3A (Study 1) and Figure 3B (Study 2).In order to account for the four different types of bilingual balance identified in The four-level categorical variables thus obtained for Study 1 and Study 2, respectively, were subsequently used as the grouping variable in two Discriminant Function Analyses (DA). DA attempts to find the best combination of predictors based on which as many cases as possible can be classified into the predetermined categories . It is u2(63) = 143.297, p < 0.001]. The first function explained 54.6% of the variance , the second explained 33.7%  while the third explained only 11.7% . The cutoff point for factor loadings was set at 0.3 (the same threshold as used for the PCA).For Study 1, three functions were identified which together significantly discriminated the four groups . The first function explained 48.6% of the variance , the second explained 34.1%  while the third explained 17.3% . Like in Study 1, Function 1 distinguished good and poor maintainers, while Function 2 distinguished good and poor learners .The DA for Study 2 also identified three functions which together significantly discriminated the four groups [Wilks’ λ = 0.143, χFigure 4B).Function 1 comprised the language of preference, the language used with friends and the self-evaluated proficiency in the L2. As in Study 1, this function seemed mainly related to success in L1 maintenance, suggesting that people who preferred the L1, used the L2 less with friends and estimated their L2 proficiency lower were better maintainers .Together, these three functions accurately predicted group membership in 76.7% of original cases , while the second investigated the development of Basic Language Cognition, in particular in relationship to lexical access  and introspective measures of proficiency and attitudes seemed to be reflected mainly in the self-ratings elicited by the Can-Do Scales in both languages. In the second analysis, findings were even more scarce and only suggested that higher levels of L1 use might have a facilitating – albeit very weak – effect on Reaction Times in lexical naming and fluency in informal speech in the L1.The somewhat disappointing results from this analysis are fully in line with previous work on L1 attrition: as was pointed out above, few studies have been able to identify any consistent impact, let alone any strong explanatory power, of predictors on actual measures of L1 proficiency and performance.For our second analysis, we therefore adopted a different approach. Firstly, we combined the different measures of proficiency  into one compound measure. This was done following Figures 3A,B, above: the area in the middle of each chart contains participants whose scores in both languages are very close to each other, but who were assigned to different groups. We feel, however, that the benefits of capturing a relationship between the two languages that may go hand in hand for some participants but be orthogonal for others outweigh these drawbacks, but we would be delighted to learn of other analyses that are able to achieve this without resorting to categorisation of data.We fully acknowledge that this classification suffers from a number of problems that categorisation of interval data invariably entails: firstly, there is substantial loss of variance incurred by collapsing all of these different scores into just four categories. Secondly and relatedly, it results in the classification of those cases who are closest to the (arbitrary) threshold established for the cutoff into one group, even though they are far more similar to individuals on the other side of the threshold in another group than to many cases in their own category. This becomes evident from the visualisation of the categorisation in Given the lack of previous insights into what factors may predict the development of a native language in immersed bilinguals, the insights gained from this classification can only be described as both unexpected and dramatic. Our hypothesis that treating language proficiency as a two-dimensional construct was confirmed by the Discriminant Analysis which was able, in both studies, to classify around three quarters of all participants accurately. Given the substantial differences between the two studies, both in terms of the population and of the linguistic skills analysed, it was particularly striking that, in both cases, the first – and hence most powerful – of the three functions identified in the analysis related to L1 maintenance (irrespective of attained level of L2 proficiency) and comprised mainly measures related to informal language use: in both studies, those participants who had higher levels of language maintenance were the ones who used the L2 less with their friends and the L1 more with their family. This finding lends support to the often intuitively held view that more informal use of the L1 should be conducive to L1 maintenance – which, however, so far has lacked empirical substantiation. For example, a study on L1 attrition of lexical access and fluency measures very similar to the ones investigated in Study 2 here and using the same set of predictors  finds noInterestingly, the first function returned by the DAs, while successfully separating good and poor maintainers, seemed unrelated in both studies with success obtained in an L2, suggesting that the amount of informal use a participant makes of both her languages does not play a strong role when it comes to the development of either HLC or BLC skills in a second language. Here, it was found across both studies that the level of education as well as the level of self-perceived proficiency seemed to play a role. In Study 1, this function also affected L1 maintenance to some extent, suggesting that, when it comes to HLC, a higher level of education may be beneficial not only for L2 acquisition but also for L1 maintenance. Similarly, in this study, the use of the L1 at work was important for both successful acquisition and successful maintenance. In Study 2, the effects of these factors were less pronounced, which is hardly surprising given that the BLC skills investigated in this study are probably much less amenable to educational levels. A rather puzzling finding is that the strongest contributing factor here was self-assessed L1 proficiency, with higher levels of proficiency being associated with better L2 skills – again, this may be related to the (unassessed) individual difference of language learning aptitude, which may have facilitated both L2 acquisition and L1 maintenance.The last function was the only one for which there was no common pattern across the two studies. In Study 1, it seemed that a more positive attitude toward the native language and its maintenance would contribute to a more balanced pattern of language dominance, while in study 2 it seemed that using either the L1 or the L2 professionally may facilitate a higher level of proficiency in both languages.Figures 5A,B depicts the median split division that was shown in Figure 3 above. However, in this case, the markers showing the position of each individual do not represent their original group membership, but the group assigned to them by the DA.It thus seems that the complex interaction of the predictors of both L1 attrition and L2 acquisition can be captured better by analyses which (a) plot out their results in a fully two-dimensional fashion and (b) do not rely on sweeping averages of the many necessary predictors, as we did through the Principal Component Analysis which yielded the independent variables used in the first set of analyses (MANCOVAs). As we pointed out above, the categorisation of data has a number of undesirable results, as it assigns cases which are very similar to each other to different groups. However, a closer look at the classifications yielded by the DA suggests that the negative impact of this may be less dramatic than one might have thought: In both studies, it is striking that, with few exceptions, most of the misclassified cases occur quite close to the median split lines. Recall that the DA does not have access to the actual scores of any of the individuals, only to the categorical group membership data. The fact that, even so, of the roughly 25% of misclassified individuals the vast majority are to be found among the more marginal cases underscores the potential value of such an analysis.Figure 5B). The participant in question is a Turkish woman who had come to the Netherlands aged 18 and, at the time of the data collection, had been living there for 27 years. While she used the L1 almost exclusively in her social life, she had in the early years of her emigration had contact with some Dutch women who had begun to teach her that language. She found that she very much enjoyed learning the language and, just before the time of data collection, had begun taking Dutch lessons for the first time (at the suggestion of her line manager). In the interview she talks about discovering aspects of the Dutch language and grammar that she had not previously been aware of, and what an enlightening and enjoyable experience this was for her. Furthermore, developing her Dutch skills also proved an empowering experience which changed her relationship to her overbearing and somewhat authoritative husband. It thus seems that, for this participant, a number of factors not measured in the present study, but probably relating to a high level of language aptitude and the experience of personal growth and self-fulfilment offered by the development of her linguistic skills, was enough to override the combination of the factors based on which the DA predicted low achievement in both languages for her.In order to gain more insight into the mechanisms of the development of language dominance, it may then be beneficial to adopt the approach suggested by Opitz, and scrutinise the more extreme cases of misclassification. We would like to illustrate this approach with the example of one individual participant in Study 2. This speaker attained the second highest score in the L1 and joint second highest in the L2, but the DA predicted her to be a poor maintainer and poor learner (represented by the purple cross toward the top right of the panel in As this and other cases in which the DA was unable to predict group membership show, the factors we included in our research design are not sufficient to paint a full picture of the circumstances under which both L1 maintenance and L2 acquisition may be more or less successful. Future studies should delve yet deeper into these questions and attempt to measure personal characteristics, such as language aptitude, and other aspects of attitude and motivation.What the study presented above shows very clearly, however, is that investigations of language dominance cannot afford to adopt a one-dimensional perspective, nor to rely on linear models of predictor-outcome relationships. We hope that these findings may inform future studies and also encourage investigations that are able to zoom in on more specific linguistic features than the relatively global and holistic ones we were able to measure here, in order to further inform our understanding of bilingual development and the forces that drive and shape it.MSS was responsible for data collection, classification and coding for Study 1, conducted all analyses reported here, and wrote the first draft of the article. GY was responsible for the data collection, classification and coding of the Turkish data in Study 2, and contributed to the revisions of the article.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.


Atomic force microscope (AFM), scanning electronic microscope (SEM) and Raman spectroscopy are used to characterize the interface of insulating substrate and graphene. The graphene films directly grown on quartz glass can attain a high transmittance of 92.8% and a low sheet resistance of 620 Ω/square. The growth mechanism is also revealed. This approach provides a highly efficient method for the direct production of graphene on insulating substrates.Synthesizing graphene at a large-scale and of high quality on insulating substrate is a prerequisite for graphene applications in electronic devices. Typically, graphene is synthesized and then transferred to the proper substrate for subsequent device preparation. However, the complicated and skilled transfer process involves some issues such as wrinkles, residual contamination and breakage of graphene films, which will greatly degrade its performance. Direct synthesis of graphene on insulating substrates without a transfer process is highly desirable for device preparation. Here, we report a simple, transfer-free method to synthesize graphene directly on insulating substrates (SiO This prominent drawback is also inevitable for the metal-CVD method, which limits its further development and reduces its superiority. The complicated transfer process of chemical etching or electrochemical bubbling involves some issues such as surface wrinkles, residual contamination and the breakage of graphene films, which will greatly degrade its performance [2/Si, Al2O3 and quartz glass by low pressure chemical vapor deposition (LP-CVD) using CO2 as oxidant and CH4 as the carbon source. The obtained graphene films show a good electrical transport property with a charge carrier mobility of 720 cm2 V−1 S−1. In addition, the direct growth of graphene on insulating substrates such as SiO2 [2O3 [Synthesizing large-scale and high quality graphene is a prerequisite for graphene applications in electronics ,2,3,4,5.formance ,27. Hencformance  directlyformance  synthesi as SiO2 ,31, Al2OiO2 [2O3 ,33, quariO2 [2O3 ,35 and BiO2 [2O3 ,37 by CVAccording to the carbon solubility in the metal substrates, the mechanisms of graphene growth on metals can be divided into two categories ,41,42,432/Si, quartz) using Cu, GO (graphene oxide), and PVA  as catalyst, seeds, and carbon sources, respectively. Atomic force microscope (AFM), scanning electron microscope (SEM) and Raman spectroscopy are used to characterize the formation process of graphene, also a tentative mechanism is presented to describe the growth processes for graphene films.Here, we report a facile method to synthesize uniform and large-area graphene directly on insulating substrates . Afterwards, the GO aqueous solution was processed by centrifuging and the supernatant were collected and used as seeds for graphene synthesis. Different sizes of GO seeds can be collected at different centrifuging speeds. GO with the size of ~260 nm is adopted in this research.0.1 g PVA powder was first added to 10 mL de-ionized water with magnetic stirring at room temperature for 2 h and was then dissolved at 90 °C for 4 h to form 1 wt % PVA solutions.2/Si wafer with 300 nm SiO2 in thickness was cleaned by ethanol, acetone, and de-ionized water respectively, then treated with Piranha solution (H2SO4:H2O2 = 3:1) at 90 °C for 30 min. Afterwards, GO seeds and PVA aqueous solution were spin-coated on the SiO2/Si wafer sequentially. Then 200 nm Cu and 100 nm CoO layers were deposited sequentially by magnetron sputter . After annealing in CVD furnace, graphene was synthesized between the Cu layer and the insulating substrate. At last, Cu and CoO layer were etched away by Marble’s reagent (CuSO4:HCl:H2O = 10 g:50 mL:50 mL), then graphene was obtained directly on SiO2 substrate without any transfer process. The process can be duplicated on other insulating substrates such as quartz glass to directly grow graphene.First, SiOThe size of GO was measured by Zeta-Size . GO aqueous solution was placed in disposable cuvettes and the dynamic scatter intensity was recorded at 25 °C. Raman spectra images of graphene films were obtained by a Jobin-Yvon HR800 Raman spectrometer with 633 nm wavelength incident laser at room temperature, spot size was 1 μm. The morphology of the products was observed by SEM (FEI Quanta-200) and an optical microscope . The thickness of the graphene was measured by atomic force microscopy (AFM)  under tapping mode with a resonance vibration frequency of 350 kHz and a scanning rate of 0.998 Hz. The electrical conductivity of graphene films was measured using a digital four-point probe system . The final result was the average of data from five devices.2/Si substrate. After treating with H2O2/H2SO4 (1/3) solution, hydroxyl groups were formed on the surface of the SiO2/Si substrate, which makes them hydrophilic. Then, GO seeds and PVA aqueous solution are spin-coated to it sequentially. 2 substrate, GO sheets and PVA are tightly absorbed to the surface of SiO2 substrate. After that, Cu and CoO layers are sequentially deposited on it by magnetron sputter. Cu layer can catalyze PVA to decompose to supply the carbon source for graphene growth and CoO layers can efficiently prevent Cu from vaporizing at high temperatures [2 substrate without any transfer process. The Energy Dispersive X-ray (EDX) test (eratures . Then grDX) test b,c shows2 substrate. The dangling bonds on the edge of the GO sheets give the marginal area high activity, so GO as seed can easily induce the deposition of carbon atoms and the growth of graphene. During the process, PVA used as a carbon source is decomposed to help the graphene grains grow up. Growth time is found to play a critical role in the morphology of graphene films grown by the present method. As shown in GO sheets with an average size of ~260 nm are adopted in this research a. Figure−1 is attributed to the first-order scattering of E2g mode, and the D band at about 1330 cm−1 arises from a breathing mode of κ-point phonons of A1g symmetry. The D band is related to the amount of disorder and its intensity reflects the degree of defects. The 2D band originates from a two phonon double resonance Raman process. Thus, the intensity ratio of D and G bands(ID/IG) of carbon material indicates the degree of the disorder such as defects, edges and ripples [D/IG decreased gradually from 1.23 to 0.31. Meanwhile the 2D (2662 cm−1) band appears, indicating that the graphene is of high quality. This may be attributed to the formation of conjugated carbon atoms and the removal of the oxygenous groups from GO. The intensity ratio of G and 2D bands (IG/I2D) as well as the full width at half maximum (FWHM) of the 2D peak were usually used to indicate the number of graphene layers. The IG/I2D value of 0.78 and 2D peak at 45 cm−1 from FWHM demonstrate the prepared graphene film was of bilayer [−1 (G band) but no peaks around 2660 cm−1 (2D bands). These results indicate that without GO seeds, PVA is decomposed and only converted to amorphous carbon. The quality of products on insulating substrates was evaluated by using Raman spectroscopy with laser excitation at 633 nm. The G band at 1570–1600 cm bilayer ,52. FiguDuring the annealing process, CoO layers can efficiently prevent Cu from vaporizing at a high temperature and promote the growth of a continuous graphene film. 2 [Based on the different refractive index between graphene and SiO2 ,55, an oTo clarify the mechanism of graphene growth with GO/PVA, in-situ X-ray absorption fine structure spectroscopy (XAFS) was used to detect the chemical state and the local structure of the copper layer during the growth process. In Due to its excellent transmittance and electrical conductivity, graphene is considered to be an ideal material to replace indium tin oxide (ITO) as transparent electrodes. However, the complex transfer process of graphene to transparent substrate usually introduces a variety of defects into the graphene sheet. The present method should provide a possible route to directly grow graphene on transparent substrates without the need for any transfer process. In this research, graphene is also directly fabricated on quartz glass. On the basis of those above-mentioned observations, a tentative mechanism is presented to describe the growth processes for graphene films, as shown in D/IG was 0.31 at 1000 °C, meanwhile the 2D band appears, indicating that the graphene is of relatively high quality. By this method, continuous graphene films can be prepared on quartz glass and used as transparent electrodes, which showed optical transmittance of 92.8% and resistances of 620 Ω/square. XAFS study shows that the growth of graphene only occurs on the Cu surface following the surface-mediated nucleation mechanism. Further work must be performed in order to evaluate the electronic transport performance of this material and therefore validate its use in practical applications like solar cells or organic light-emitting diodes.In summary, we have developed a transfer-free method for the large-scale synthesis of graphene on insulating substrate using a Cu layer, GO and PVA as the catalyst, seeds and carbon sources, respectively. During the growth process, GO seeds can act as the nucleus to efficiently help the graphene films to form. Raman spectra shows a ratio of I


Programmed death 1 (PD-1), when activated by its ligands PD-L1 and PD-L2, suppresses active immune cells in normal immune regulation to limit autoimmunity and, in tumors, as a mechanism of immune evasion. PD-L1 expression has been described as both a prognostic and predictive marker in many solid and hematologic neoplasms, as targeted therapies against the PD-1/PD-L1 interaction have gained clinical importance. PD-L1 expression has been assessed in a few studies on mastocytosis. We review this literature and the need for further investigation of the tumor-immune interaction in mastocytosis. Programmed death 1 (PD-1), first described in 1992, has been described on many activated immune cells, including both CD4 T-cells, CD8 T-cells, B-cells, NK/T cells, dendritic cells and macrophages. When PD-1 binds its ligands, PD-L1 and PD-L2, active immune cells are inhibited as an important check on protecting tissues from autoimmunity [PD-L1 expression has been assessed in many solid and hematologic malignancies, both as a prognostic marker and as a predictive marker of the response to antibodies that block the PD-1/PD-L1 interaction ,3,4,5. PAn immunohistochemical (IHC) assessment of PD-L1 expression has been used to predict the response to and qualify patients for immune checkpoint inhibitors . In someBecause PD-L1 expression in tumor cells represents a mechanism of immune evasion via the interaction between the PD-L1 ligands on tumor cells and PD-1 on lymphocytes to suppress the immune response, an assessment of PD-1 within tumor-infiltrating lymphocytes is important for understanding the tumor-immune interaction. In melanoma, previous studies have examined the role of both innate and adaptive immunity, where PD-L1 expression has been proposed in some cases to be driven by oncogenic mutations, and in others, to be mediated by cytokines . While patients with cutaneous mastocytosis (CM) and indolent systemic mastocytosis (ISM) generally do not require anti-neoplastic therapy, current therapeutic options in advanced systemic mastocytosis (advSM) include cladribine, midostaurin, imatinib, and hematopoietic stem cell transplants . MultiplKIT mutations. Following stimulation of the mast cell lines with recombinant PD-L1, there was suppressed growth within the LAD2 cell line. In 2013, Kataoka et al. assessed the PD-1 expression by IHC within skin biopsies from patients with CM compared to patients with non-specific dermatitis and demonstrated PD-1 positivity within 10/30 CM cases and no PD-1 staining within the non-specific dermatitis cases [In 2016, Kuklinski et al. assessed the PD-L1 expression within 16 skin biopsies of patients with diverse mastocytosis types, including diffuse CM, solitary mastocytoma, and SM involvement (mastocytosis in skin) and found strong and diffuse PD-L1 staining within all mastocytosis specimens, regardless of the subtype . There wAdditionally, in 2016, Rabenhorst et al. examined PD-1, PD-L1, PD-L2, and tryptase within the serum of 43 patients with mastocytosis and 22 healthy controls, demonstrating significantly increased PD-L1 within the serum of adult patients with mastocytosis, compared to controls. In addition, serum PD-L1 was significantly elevated in patients with aggressive SM (ASM), SM with an associated hematologic neoplasm (SM-AHN), and mast cell leukemia (MCL), compared to patients with a non-advanced disease . They foIn 2018, Hatch et al examined PD-1 and PD-L1 by IHC in 55 tissues from patients with mastocytosis, demonstrating a PD-L1 expression in 77% of bone marrow biopsies and 92% of skin biopsies including all mastocytosis types; they found no expression of PD-1 or PD-L1 within mast cells in healthy or reactive bone marrows or cases of myelodysplastic syndrome or myeloproliferative neoplasms . PD-L1 sSeveral groups have demonstrated PD-L1 expression in neoplastic mast cells. While PD-1 expression was present in a subset of CM cases reported by both Katoaka et al. and Hatch et al., interestingly, PD-L1 expression was limited to neoplastic mast cells and was not reported in tumor infiltrating lymphocytes ,16. The In the Rabenhorst et al. 2016 cohort, there was a breakpoint with significantly higher PD-L1 in the serum of adult patients with advSM , compared to non-advanced mastocytosis (ISM and CM). There was no clear difference in the intensity of the staining or the percentage of the mast cells staining for PD-L1 by mastocytosis subtype by IHC in the Hatch et al. cases. While the prognostic and predictive value of PD-L1 expression within tumor tissue is well-established, the clinical utility of serum PD-L1 measurements is under investigation and is still unknown, limited by a lack of robust data .In addition to the difference in the PD-L1 detection by mastocytosis subtype in serum versus by IHC on tissue biopsies, there was a variability seen in the percentage of mast cells staining for PD-L1 within the same splenic tissue reported in Hatch et al. This variability of staining within the same tissue has implications for the use of PD-L1 as a potential prognostic or predictive biomarker. Future studies could include both a serum and tissue assessment of PD-L1 with an assessment of PD-L1 in multiple tissues within the same patient, in order to better characterize the expression within different types of mastocytosis and the variability in the staining patterns. The PD-L1 expression in neoplastic mast cells that has been confirmed by these studies warrants further investigation of the tumor-immune interactions in mastocytosis to clarify possible breakpoints in the PD-L1 expression by type of mastocytosis, as well as to clarify the role of the immune checkpoint molecule expression in tumor infiltrating lymphocytes, and to explore a possible role for immune checkpoint inhibition.