The paper presents the results of research on the use of flake graphene as an additive to plastic grease in order to improve its tribological properties. The influence of concentration (0.25–5.00 wt.%) and the form of graphene  on selected properties of the base grease were investigated. It has been found that the addition of graphene flakes improves the anti-wear properties of the lubricant. The greatest improvement in the properties of the lubricant was achieved by using graphene at a concentration of 4.00 wt.%; the reduction in the average diameter of the wear scar was almost 70% for GO and RGO, compared to the base lubricant without the addition of graphene. The factor driving the growth of the market for these products in the world is the dynamic technological development. Lubricant manufacturers have to keep pace with more and more advanced vehicles as well as innovative machines and devices, where precision and smooth operation are crucial for their efficient and profitable operation. During mechanical processes, as a result of strong friction between the surfaces, wear of the mechanical elements of the friction junction occurs and heat is generated, which ultimately reduces the efficiency of work and shortens the life of the device. It is estimated that about 23% 119 EJ) of global energy consumption is due to anti-friction and regeneration of friction elements [19 EJ of missions . Therefomissions ,6.Flake graphene, which is a known 2D carbon nanomaterial, is obtained from graphite. At the same time, it should be emphasized that graphite is characterized with greasing properties and it is widely implemented, i.e., as an anti-wear agent. Graphene is another allotropic type of carbon, next to graphite. GO is the oxidized form of graphene, decorated with oxygen functional groups . GO particles are highly hydrophilic and form stable dispersions in water and some organic solvents. RGO is obtained by reduction of oxygen functional groups of GO. Contrary to GO, rGO is hydrophobic and show high mechanical strength, large specific surface area and high electric and thermal conductivity .The use of flake graphene is currently the subject of much scientific research in various areas, such as composites , functioAs part of the research described in the literature, the tribological properties of various lubricants  with the addition of various forms of graphene flakes  were analyzed. The tribological properties of polyalphaolefin-2 (PAO2) base oil with the addition of multilayer graphene containing 0.05 wt.%, 0.10 wt.% and 0.50 wt.% were investigated using a 4-ball tester. The tests were carried out at loads from 120 N to 400 N and rotational speeds in the range of 100–400 rpm . It was Graphene as an anti-wear additive to plastic greases was also tested. The properties of mineral plastic grease thickened with lithium saponified, enriched with the addition of graphene nanoplatelets with a concentration of 0.50 wt.%, 1.00 wt.%, 5.00 wt.% and 10.00 wt.% were investigated. Various loads were applied: 5 N, 10 N and 15 N . It was Flake graphene offers great research opportunities because its surface can be chemically modified and its properties can be controlled in this way. For example, the lubricating properties of monolayer graphene oxide when used as an additive to water-based lubricants were investigated . TriboloBased on the literature review, it can be concluded that the addition of graphene to lubricants improves their tribological properties. However, graphene still remains the object of tribological research and the obtained results are not always unequivocal. This may result from the use of various forms of graphene , different sizes of flakes, different number of carbon layers, different content of oxygen functional groups on the surface and different media in which graphene and its derivatives are dispersed, such as also different test conditions. Therefore, the available information is still incomplete and requires further research. The complexity of the application of graphene in terms of improving the tribological properties of lubricants is also indicated by Larrson et al. . MoreoveBearing in mind the above-mentioned limitations of the available literature data, this work proposes an innovative approach to the implementation of graphene materials as a nanoadditive with anti-wear properties to greases. For the purposes of the work, graphene materials were designed and manufactured, representing two types of flake graphene: graphene oxide and reduced graphene oxide. The paper presents the comparative results obtained for both materials. In order to obtain the best knowledge about the influence of graphene materials on the anti-wear properties of the grease, the tests were carried out for a wide range of concentrations of graphene materials in the grease, amounting to 0.25–5.00 wt.%. The experiment designed in this way allowed to obtain a detailed quantitative description of the issue under study. In addition, the article attempts to answer which of the two tested types of graphene materials will better meet the requirements for carbon nanoadditives for lubricants, taking into account their practical application.® —The research was conducted using graphene oxide powder and reduced graphene oxide powder by G-Flake® CrossBeam® Workstation, Carl Zeiss, Oberkochen, Germany, at a voltage of 1 kV, energy selective backscattered EsB detector), Raman spectroscopy , elemental analysis . Regarding the SEM experiment, it was implemented in order to visualize GO and RGO materials. The powders were placed on a silicon wafer and then glued with a carbon tape to an SEM table. No coating with conductive agent was conducted prior to visualization. As for the Raman examination, collected raw data were analyzed with Wire 3.54 software provided by the spectrometer producer. The analysis was carried out in order to discriminate the position of D and G peaks using a Gaussian/Lorentzian. Prior to determination of the peaks, position baseline subtraction was implemented. Detailed characterization of the purified GO water suspension, from which GO powder and RGO powder were then obtained, is presented in our earlier publication [Graphene oxide was produced using a modified Hummers method. This method is based on the oxidation of flake graphite  in sulfuric acid VI  with the addition of potassium permanganate  and potassium nitrate . After the oxidation process, the graphite oxide in the form of an aqueous suspension is exfoliated and then purified by filtration. After the purification process, an aqueous suspension of graphene oxide is obtained. In order to produce reduced graphene oxide, graphene oxide undergoes a reduction reaction, which takes place at high a temperature in the presence of a reducing agent—hydrazine . After the reduction process, the reduced graphene oxide is purified by filtration. To obtain GO or RGO in powder form, the aqueous material dispersion was dried. The obtained graphene materials (GO and RGO) were then characterized using the following methods: scanning electron microscopy (SEM)  and lithium stearate  thickener. It was prepared in laboratory conditions by adding the thickener lithium stearate to the paraffinic base oil, mixing and then heating with constant stirring until the thickener completely dissolved (to approx. 220 °C). The dissolved mixture was allowed to cool completely at room temperature.The lubricant compositions were prepared in laboratory conditions by adding the appropriate amount of GO or RGO in the form of a powder to the lubricant base. The samples were then mixed using a planetary mixer  to obtain a homogeneous dispersion . Mixtures of grease and flake graphene were subjected to tribological tests, in which the concentration of graphene  was from 0.25 wt.% up to 5.00 wt.%. It was observed that as a result of the shear stresses arising when mixing pure base grease and base grease with the addition of graphene, the viscosity of the grease decreased due to the shear-thinning phenomenon .The authors of the study  describe3). These balls are pressed down with a ring and fixed with a tightened clamping nut. The fourth ball (movable) is mounted in the upper holder. During the test, it rotates at a speed of 1450 ± 50 rpm, which corresponds to the speed of the rubbing surfaces of 0.55 m·s−1. The balls in the lower holder are pressed with the force P against the ball mounted in the upper holder by means of a lever. The friction linkage diagram is shown in The tests of the lubricating properties of the obtained plastic greases with the addition of flake graphene were determined by measuring the average diameter of the wear scar. The tests were performed with the use of a 4-ball tester . In this tester, the elements of the tribological couple are in point contact and a sliding movement is performed. The tribological couple in the tester consists of four balls with a diameter of 12.7 mm, made of bearing steel with a hardness of 62.7 HRC. Three balls (stationary) are placed in the lower holder, into which the tested lubricant is also introduced  results from the presence of vacancies or dislocations in the graphene layer and at the edge of this layer. This band is also related to the presence of defects in the material. The G band (located near 1590 cm−1) corresponds to the sp2 hybridization of the carbon network and is attributed to the first-order scattering from the doubly degenerate E2g phonon modes of graphite in the Brillouin zone center as well as bond stretching of sp2 carbon pairs in both rings and chains [−1 for GO and ~1347 cm−1 for RGO, respectively, and a G peak at ~1586 cm−1. The ID/IG ratio for GO is 1.05 and for RGO 1.20. The value of the ID/IG parameter increases with the increase in the degree of disorder in the structure of the graphene material.d chains . The ID/d chains . The anaThe results of elemental analysis of graphene materials are presented in The grease is a colloidal system, where the dispersing phase is oil and the dispersed phase is thickeners. The role of the thickener is to ensure the proper spatial structure of the plastic grease and the specific, required rheological and tribological properties. The spatial structure formed in the lubricant by the thickener fibers, as a result of the action of shear forces and the temperature increase during the operation of the tribological node, disintegrates ,32. An ePrepared lubricants with different contents of graphene material  were subjected to anti-wear tests. In the case of lubricant samples containing the additive in the form of RGO, the trend observed was similar to the compositions with the addition of GO. In this case, an increase in the wear of the test elements by 24% (4.03 ± 0.27 mm) was also observed compared to the base lubricant (3.26 ± 0.28 mm) for the lowest additive content of 0.25 wt.%. Increasing the concentration of graphene to 0.50 wt.% results in a wear scar with an average diameter close to the scar obtained when lubricating the node with base grease without additive. Only higher concentrations of the nanoadditive result in a significant reduction in consumption. The lowest wear was observed at 4.00 wt.%. RGO concentration (scar reduction by approx. 70%). Increase in RGO concentration in the lubricant from 1.00 to 4.00 wt.% reduces the average diameter of the wear scar from 2.51 ± 0.23 mm (23%) to 1.01 ± 0.05 mm (69%), respectively, compared to the grease without additive. Further increase of the additive content to 5.00 wt.% may increase the wear of the balls (1.97 ± 0.09 mm). It can be concluded that the RGO concentration of 4.00 wt.% in the grease is the limit concentration, exceeding which the antiwear properties of the grease may deteriorate. Even though 5.00 wt.% RGO concentration increases the consumption in relation to the concentration of 4.00 wt.%, in relation to the grease without the addition of graphene, the average diameter of the wear scar was reduced by 40%. Deterioration of lubricating properties for the highest concentration of the additive 5.00 wt.% has also been observed as a result of the use of GO. In the case of high concentrations of graphene, agglomerates form in the lubricant, which negatively affect the friction surface protection. The resulting tendency to change the wear properties of a lubricant with an increase in graphene concentration is consistent with the results of the research carried out by Fu et al. . AccordiDuring the tests of lubricants, the color of the lubricant with the addition of GO changed from brown to clearly darker, which may suggest that the graphene oxide is reduced during friction ,38. The Tests were also carried out on lubricants with the addition of RGO for a higher load of 100 kgf for 1 min and compared with the results obtained in the tests with a load of 60 kgf and a duration of 60 min, and the results of the measurements are presented in Grease samples were produced with the addition of GO and RGO with a concentration in the range of 0.25–5.00 wt.%. It was found that the addition of flake graphene improves the anti-wear properties of the lubricant, while the concentration and type of graphene have an impact on the tested lubricant properties and its stability. For lower contents of graphene additive (0.25 wt.% and 0.50 wt.% for GO and 0.25 wt.% for RGO), an increase in wear is observed as compared to the base grease. For low RGO concentrations (0.50 wt.%), the grease properties are similar to those of the base grease without additive. The introduction of flake graphene to the base of the plastic grease may disturb the internal network of the grease due to the high activity of the surface of the graphene flakes. This phenomenon may be the cause of increased node wear at low additive concentrations (0.25 wt.% and 0.50 wt.%), as these contents are probably too low to compensate for the breakdown of the lubricant’s inner network. This effect is particularly evident in GO containing numerous oxygen functional groups on the surface of the flakes.The use of a higher concentration of graphene 1.00–5.00 wt.% results in a reduction in the average diameter of the wear scar, which is greatest for an additive content of 4.00 wt.% . A further increase in the concentration of the additive to 5.00 wt.% results in a renewed increase in wear, which may result from the agglomeration of the graphene material and a significant increase in viscosity, so that the lubricant is not able to produce a lubricating film that will protect the friction surface well.Out of the two tested forms of flake graphene, RGO is a recommended additive for lubricants due to its stability under operating conditions, while GO was reduced during friction. The RGO powder tested in operation can therefore be successfully used as a material improving the tribological properties of lubricants.The use of flake graphene in lubricants is not a thoroughly researched and explained issue. In order to further develop the application of graphene flakes as a lubricant additive, it is necessary to continue research, including in the direction of chemical modification of the flake surface, and to explain the mechanisms of graphene operation during friction, and also to maintain the properties of lubricants unchanged in subsequent work cycles.


A sheet resistance value as low as 100 Ω/□ is reported for a 3-layer stacked graphene:N film.Nitrogen substitutional doping in the π-basal plane of graphene has been used to modulate the material properties and in particular the transition from hole to electron conduction, thus enlarging the field of potential applications. Depending on the doping procedure, nitrogen moieties mainly include graphitic-N, combined with pyrrolic-N and pyridinic-N. However, pyridine and pyrrole configurations of nitrogen are predominantly introduced in monolayer graphene:N lattice as prepared by CVD. In this study, we investigate the possibility of employing pyridinic-nitrogen as a reactive site as well as activate a reactive center at the adjacent carbon atoms in the functionalized C–N bonds, for additional post reaction like oxidation. Furthermore, the photocatalytic activity of the graphene:N surface in the production of singlet oxygen ( Nowadays, however, the good reputation of graphene as a promising material for various applications11, is also due to the possibility to modulate its chemical and physical properties through chemical functionalization and, in particular, doping16.Since the publication on graphene by Geim and Novoselov in 2004, and after more than one hundred thousand publications, there is no doubt that the success of graphene lies in its fascinating properties such as one-atom thickness, transparency, thermally and electrically conductivity, high mobility of electrons and flexibility14. Specifically, when nitrogen atoms are substitutionally incorporated into the π-lattice of the carbon ring, they add resonant electrons, which provide n-doped graphene sheets. Nevertheless, the landscape of the nitrogen functional forms possibly present in nitrogen-doped graphenic materials is very rich19. In CVD nitrogen-doped graphene, depending on the doping procedure, the nitrogen moieties include graphitic-N, combined with pyrrolic and pyridinic nitrogen18. Among these bonding configurations, graphitic-N induces an n-type doping effect, whereas pyrrolic-N and pyridinic-N may result in either weak n-type or p-type doping20. To date, there are quite a number of studies focused on the growth of N-doped graphene by direct or post treatment processes, which show the capability of tuning graphene chemical and physical properties25. However, none of them has reported that the nitrogen functionalities inserted in the basal plane of the carbon lattice may be further modified to further improve the conductivity of the layer. Thus, from a chemical standpoint, N-doped graphene is an appealing graphene type, because nitrogenous functionalities and, in particular, pyridine nitrogen can be a reactive center as well as activate a reactive center at the adjacent carbon atoms in the functionalized C-N bonds for additional post reaction like oxidation27. In addition, the presence of these N-functionalities in the graphene structure, besides changing the carrier density , gives an interesting catalytic activity of the graphene surface, such as the well investigated activated generation of reactive oxygen species (ROS)31. Among these new “catalytic” capabilities, the production of singlet oxygen (1O2) by both (i) activating the dissociation of peroxydisulfate28 and (ii) the photosensitize excitation of molecular oxygen30 is of particular interest. In fact, singlet oxygen ), being a non-radical reactive oxidizing specie, does not intervene on the conjugated double bond as it happens for radical species  and shows higher reactive selectivity also because of its electrophilic nature.Graphene doping can be performed through two methodologies: (i) surface chemical functionalization, both covalent  and non-covalent  and (ii) substitutional doping in the hexagonal lattice of the graphene with heteroatoms, mainly nitrogen and boron, capable to control the n-type and p-type character of the graphene, respectively2 graphene network. As for the CVD growth of N-doped graphene on copper foil, the major challenges are how to control the insertion of the different N-functionalities. Specifically, in order to realize an increase in p-type doping, we need to minimize the graphitic nitrogen in favour of pyrrolic and, even better, pyridinic nitrogen; this is because each nitrogen substituting a graphitic carbon increases the number of conjugated π-electrons, thus improving the n-type conductivity. It is important to underline that N-graphitic atom, like carbon atom in pure graphene, has a low reactivity. Among the different methodologies proposed in literature, we opted for the CVD growth of N-functionalized graphene (graphene:N) via N atoms embedded into Cu substrate and used as a nitrogen solid source24. We show that this approach allows obtaining N-graphene layers with small grain size and therefore a high density of structural defects, such as grain boundaries and carbon vacancies, which are the preferred sites for the inclusion of pyridinic- and pyrrolic-nitrogen. From this point of view, in other words, graphene grain-boundary defects do not damage the material properties but rather improve them32. The post-growth treatment of N-doped graphene to realize chemical modifications at nitrogen reactive sites is performed by photoirradiation with Xe lamp of N-graphene sample in air. The chemistry, involving the interaction of singlet oxygen photo-catalytically activated on N-graphene, is highly selective towards pyridine active sites leaving the basal plane components intact. We show that the pyridine ring almost disappear in favour of pyridine N-oxide and pyridone structures, both having a strong p-doping effect.In this paper, we report on N-doped graphene, grown by CVD, and its subsequent chemical modification to improve the p-type conductivity, taking advantage of the presence of nitrogen functionalities as new active sites for selective reactions, without affecting the spare there possibilities to functionalize the aryl carbon in such a way as to generate new electron-withdrawing groups, thus improving p-doping? The outcomes arising from the present study confirm positively the hypothesis we set out. Figure +-doped graphene with some pyridinic groups converted to pyridine N-oxy and pyridone groups.To enhance the performances of graphene employed as a transparent conductor, a uniquely accessible route is to reduce the sheet resistance by operating a chemical doping. Therefore, it is essential to fingerprint the structure of the graphene layer and to understand the possible chemical modification for achieving high efficiency and stability in the charge transport process. For the nitrogen doped graphene (G:N), since the presence of C-N hetero-bonds into the aromatic platform, the main question is: 3 defects in the C-sp2 basal-plane. Here, it is important to underline that Pyridinic-N and Pyrrolic-N occur mainly at the boundary of C-vacancy sites or, even more, at the edge of the graphene grains19. Therefore, since the Pyridinic-N is responsible for the activation of reactive sites leading to p+-doping, a higher defects density  in G:N introduced during the growth allows for higher p-doping33.Looking ahead, from the side of the growth and the doping chemistries, both Raman and XPS data confirm (provided evidence) that graphene (G:N) incorporates N atoms during the growth and contains the “new” N-functionalities without introducing significantly C-sp35.Figure 13. The size of graphene grains is around 10 µm. Two typical spectra with G (∼1582 cm-1) and 2D (∼2718 cm-1) peaks are observed without apparent D (∼1350 cm-1) peaks indicating the high quality of the graphene layer. The ratio I(2D)/I(G) = 1.8 and the G and 2D peak bandwidths of, respectively, 14 cm-1 and 27 cm-1 are fingerprint of a single layer graphene (blue line). The Raman spectrum with I(2D)/I(G) ∼1 (red line), recorded at the grain centre, is representative of bilayer. Most of the area is covered with single layer graphene, whereas, the presence of the small double layer island in centre of each grain confirms the under-layer nucleation mechanism described by Nie et al.36 The distribution density of double layer islands on graphene can be evidenced by enhanced optical contrast of the microscope image of graphene transferred on 300 nm SiO2/Si reported in Figure S1 together with the relative Raman analysis.Optical microscopy and Scanning Raman spectroscopy were used for quality mapping the graphene layer Fig. c. The mi13. Besides the small contribute of carbon solubilized in copper foil (peak 1 at 283.6 eV)37, there are no significant changes in the deconvolution going through the three samples. The assignment of the peak deconvolution indicates that the largest proportion of C-functionalities are always associated with presence of sp2 carbon with C = C bonds (284.4 eV) and carbon–oxygen as follows: C–OH (285.2 eV), C–O–C (286.9 eV), C = O (287.9), O = C–O (288.8 eV). The presence of carbon-to-nitrogen bonds adds a new peak at 286.0 eV ascribed to N = C bonds that are located in the π-conjugated graphene structure39. On the other side, significant changes are observed on N1s peak deconvolution.Figure 41, it is recognized the presence of the three typical components of pyridinic , pyrrolic  and graphitic/quaternary  nitrogen bonds40. The peak of graphitic nitrogen (Ngr) is much lower than pyridinic (Np ) and pyrrolic (Npr) nitrogen, thus the n-type doping with two p-electrons at the graphene π-cloud is low. Almost equivalent peak intensities are observed for Np and Npr signals for as grown graphene on copper foil. Following the graphene:N single layer transfer on Si/SiO2 substrate and the subsequent Xe-lamp irradiation, the N1s peak deconvolution needs two additional contributions at 400.7 eV and 402.3 eV assigned to Pyridone (Npd) and Pyridine N-oxide (Nox), respectively38. The appearance of these new N-functionalities takes place at the expense of pyridine nitrogen as evidenced by the strong reduction of the Np energy peak, while Npr peak, and, thus, the amount of pyrrolic nitrogen structures remains unchanged.Specifically, in the pristine graphene:N (G:N) grown on Cu substrate, besides the contribute at high binding energy (404.6 eV) due to adsorbed nitrogen42. The proposed chemistry is outlined in the following overall reactions (Eqs. (1O2). Here, graphene:N surface is the catalyst that activate the hydrolysis of ammonium persulfate for the formation of superoxide radicals. Specifically, mainly pyrrole nitrogen acts on the neighbouring carbon atoms by increasing the charge density, thus generating an adsorption and activation center to form singlet oxygen42.The observed evolution of the N1s peaks can be read in the chemical processes involving the interaction of G:N with singlet oxygen. In fact, the formation of singlet oxygen occurs on pristine G:N during both (a) the solubilisation of cupper foil by ammonium peroxydisulfate and (b) the light irradiation by Xe-lamp. As for the generation of single oxygen by interaction of G:N layer (still on TRT) with ammonium peroxydisulfate during the phase of copper solubilisation, the chemistry has been described in detail in previous studies29 have evaluated the role of graphene functionalization in determining the ROS generation. In particular, they underline that the generation of 1O2 is driven by graphene photoexcitation with energy larger than the excitation energy of O2-ground state (3Σg-) to 1O2 (1Δg), which is of approximately 0.97 eV. This photosensitization mechanism has been reported as the most common means of singlet oxygen generation43. It is detailed that the energy transfer to O2 (3Σg-) from an excited state of a sensitizer, which is formed by the light absorption of light in a specific wavelength region, results in the formation of both excited states O2(1Σg-) and 1O2 (1Δg). And, followed by the very fast spin-allowed 1Σg- → 1Δg deactivation that results in the complete formation of 1O2 (1Δg). Moreover, it has been demonstrated that the graphene doping by heteroatoms, e.g., nitrogen, increase the photocatalytic efficiency44. The photochemical production mechanism of singlet oxygen on graphene:N can be depicted as follows:The graphene:N shows its catalytic action also in the photogeneration of reactive oxygen species (ROS) and, in particular, of singlet oxygen by molecular oxygen. In a recent paper, Yao and coworkers1O2-treatment process is carried out also on substrate-transferred graphene:N (glass or SiO2/Si substrates) by irradiation with Xe lamp. The effectiveness of the photocatalytic aerobic oxidation via singlet oxygen has been exploited by many for the realization of selective oxidative processes towards organic and biological molecules30.Thus, the completion of the 46. In particular, since the high lifetime of 1O2 in the gas phase, reducing singlet oxygen is expected to facilitate the first electron transfer in ORR. The general scheme for the complete reduction of O2/1O2 molecules to OH- is as follows:Furthermore, nitrogen-doped graphene is an active catalyst also in the oxygen reduction reaction (ORR), in which again the singlet oxygen can play an important role27, as schematized in Fig. Nevertheless, the same oxidative processes can occur on the reactive sites present on the graphene surface; among these, the pyridine site is important as it can be oxidized simultaneously by aerobic photoirradiation to N-oxy pyridine and pyridineNp at 398.8 eV) is in favour of N-oxy pyridine (Nox at 402.3 eV) and hydroxyl-pyridone (Npd at 400.7 eV). While the other two pyridone-tautomers (carbonyl and zwitterion) with a protonated nitrogen, which resemble a quaternary nitrogen with a very similar XPS peak position to the graphitic nitrogen, contribute to the observed increased of the peak Ngr at 401.5 eV. The scheme emphasizes the role of the delocalization of the positive charge on the aromatic ring in both the oxidation to N-oxide pyridine and the formation of pyridone by the attachment of the OH group to the carbon atom in α-position to the pyridine nitrogen. The C1s XPS spectra shows that the C–OH, peak 3 at 285.5 eV, slightly increases after transferring on substrates. This confirms that carbon atoms close to pyridine nitrogen are the main active sites, among the different doping configurations of nitrogen, capable of further functionalizing graphene in the direction of p-doping27. Thus, the possibility of maximizing the introduction of N heteroatoms with pyridinic structure in the sp2 carbon lattice would maximize the p-doping effect.The observed changes in nitrogen configurations, as assigned in Fig. 2. The observed slow kinetics is consistent with complex formation pathway of singlet oxygen, whose concentration is determined not only from light irradiation  measurement, i.e. the shift of the Dirac point at high positive voltage (VD = 39 Volts) for the light irradiated graphene:N when compared with pristine graphene (VD = 20 Volts). The evaluation of hole carrier mobility has been done using the formula µh = (L/W)·C-1·VD-1·(dID/dVG), where C = 120 µF·m-2 is the gate capacitance of the silicon dioxide dielectric layer (300 nm thick)49. The measured hole mobility of annealed pristine-G and light irradiated graphene:N are nearly equal and reach an average values of about 1350 cm2/(V·sec). Thus, the reduction of sheet resistance in irradiated graphene:N is mainly ascribed to the increase of hole carrier density.The changes in G:N surface structure in the direction of a p-doping have been monitored by Raman spectroscopy and electrical characterization, both sheet resistance (Van der Pauw) and FET measurements. Figure  see Eq.  in the p-1, from 1580 cm-1 to 1604 cm-1; (iii) the G peak asymmetry and Lorentzian-deconvolution in two narrow peaks (G1 at 1596 cm-1 and G2 at 1604 cm-1) after Xe-lamp irradiation; and (iv) the high-energy displacement of the 2D peak that moves of 9 cm-1, from 2675 cm-1 to 2684 cm-1. Similar results are observed from the Raman analysis of graphene:N transferred on glass before and after Xe lamp irradiation  is confirmed by the Raman analysis of single layer graphene on FET device. Figure n Figure .Figure 648. The observed asymmetry of the G-peak indicates the existence of charge inhomogeneity within the laser probe, i.e., on a scale of 1 µm50.The simultaneous occurrence of these four events can be read in the more general picture of a strong increase in p-type doping (increase in the carrier density of holes) as described in the Raman "master paper" on the doping effect by Das et al.49, from these important Raman parameters we can estimate an hole doping density of about 1.3 × 1013 cm-2 for the irradiated graphene:N sample. This value is consistent with the average value of sheet resistance measured for a single layer of G:N after irradiation. In fact, by applying the simple Drude model of electrical conductivity, Rs (Ω/□) = 1/(n·µ·e), where n = 1.3 × 1013 (cm-2), µ = 1350 (cm2V-1 s-1) and e = 1.6 × 10–19 (A·sec), we extract the sheet resistance of 350 (Ω/□) that matches the measured values shown in Fig. According to ref.13. Figure 13.Multilayer graphene samples on Corning glass substrates have been prepared by using the layer-by-layer procedure described in ref.In summary, using a Nitrogen-doped graphene deposited from a direct CVD method, an efficient p-doping effect has been demonstrated through the selective photochemical activation of pyridinic nitrogen to N-oxy pyridine and pyridone. Notably, the sheet resistance of light-irradiated Nitrogen-doped graphene significantly reduces and can even reach a value as low as 100 Ω/□ for a three-layer graphene.3, SOCl2,…) for the covalent doping or it is required to host molecules or particles for the non-covalent doping, the photochemical doping of graphene:N has the advantage to be easily applicable to any combined graphene-semiconductor device architecture. In addition to the capability of doping graphene on any surface device, this light-driven doping strategy has great potential to realize selective-area doping by photolithography mask or laser patterning.Unlike conventional chemical doping, where the graphene layer is submitted to chemical reaction with strong oxidants (HNO2 size) was inserted in a quartz tube of the thermal-furnace CVD reactor. The quartz-tube was evacuated to a vacuum better than 10-3 torr and heated to 100 °C under an N2 gas flow of 200 sccm (0.2 torr) that was maintained for 20 min after the temperature was stabilized. The copper nitridation process was carried out in the nitrogen plasma downstream under the following conditions: N2 flow rate 200 sccm, pressure 0.2 torr, rf (13.56 MHz) power 250 watts, exposure time 30 min. Subsequently, the temperature was raised to 990 °C under an H2 gas flow of 10 sccm (0.05 torr) that was maintained for 20 min after the temperature was stabilized. The annealing process was followed by the graphene growth: CH4 (5 sccm) was added to the gas feed for a growth time of 20 min. After the growth phase, the furnace was moved from the growth-zone to realize the rapid cooling of the graphene/copper foil.The growth of nitrogen-doped graphene occurs in two phases: the plasma nitridation of copper foil and the catalytic CVD-growth. Both processes were carried out in the same quartz tube (i.d.10 cm) equipped with external capacitive-coupled electrodes for the plasma nitridation process and furnace for the CVD growth. The copper foil  using the thermal release tape (TR-tape) procedure. The tape was placed on top of the graphene/copper foil and pressed by the laminator. The copper was etched away in an ammonium peroxydisulfate, ((NH4)2S2O8) solution (20 gr/liter) and the floating sheet of graphene/TR-tape was rinsed in DI water and air-dried. Hot-pressing (at ≈100°C) performed the graphene transfer onto substrates, thus the tape was then peeled off. Multilayer graphene samples on Corning glass for sheet resistance measurements were fabricated by the layer-by-layer transfer procedure described in ref.13.The graphene layer was then transferred to the substrate  with maximum wavelength emission at 470 nm. During light irradiation, the sheet resistance was monitored in real time to ensure the completion of the variation process.. The laser power was kept at 1.0 mW to avoid laser-induced heating effect.Raman spectra of the pristine graphene:N layers and of the photochemical modified layers were recorded with a LabRam HR (Horiba JY) system. Spectra on glass and SiO2/Si were measured at room temperature with 532 nm laser light and captured using 100x objective lens magnification with a focusing laser spot less than 1µm in diameter. Spectra of graphene on copper foil were collected using 473 nm laser2 graphitic component of the C1s spectrum (284.4 eV) was used as internal standard for charging correction. The high-resolution spectra were fitted with mixed Gaussian-Lorentzian peaks after a Shirley background subtraction. The standard deviation in the peak position was ± 0.1 eV.The surface chemical composition of doped samples was investigated by X-ray Photoelectron Spectroscopy (XPS), using a Theta Probe spectrometer (Thermo VG Scientific) equipped with a monochromatic Al Kα X-ray source (1486.6 eV) operated at 15 kV and a spot size of 400 µm, corresponding to a power of 70 W. Survey (0–1200 eV) and high-resolution  spectra were recorded in FAT  mode at pass energy of 150 and 50 eV, respectively. All spectra were acquired at a take-off angle of 37° with respect to the sample surface. A flood gun was used to balance the surface charging. The C1s signal for the spA Keithley 2400 Source Meter was used to monitor the electrical properties of graphene layers. Sheet resistance was measured in the 4-probe Van der Pauw configuration.2 / Si substrates  were used for the field effect measurements. The FET device was fabricated by gold evaporation of the source and drain contacts . Measurements were performed in air and in vacuum (better than 10-3 torr) with a Keithley 2400 analyzer at room temperature. A back-gate bias (Vg) ranging from −80 to +110 Volts was applied on the Si side of the SiO2/Si substrate. Before measurement, the FET device was kept under vacuum overnight and treated by thermal annealing @ 200 °C. This allows to desorb surface species on graphene.Single layer graphene on 300 nm SiOSupplementary Information.


Meanwhile, no evidence of malignancy was found, and pathological manifestations led to the subsequent six months of anti-tuberculosis treatment. Combined with the fact that standard anti-tuberculosis showed no improvement, and the patient’s condition was stabilized by corticosteroid treatment alone, a final diagnosis of sarcoidosis was made by an MDT (multidisciplinary consultation). Reported herein is the first case of a hyper vascularization condition within the non-typical asymmetrical sarcoidosis lesions, which should help to establish that the uptake of 3PRDG2 in sarcoidosis can avoid imaging pitfalls.Sarcoidosis is a multi-system disease of unknown etiology that typically occurs in middle-aged adults, often presenting as the formation of granulomas in various organs, including the lungs. Non-typical pulmonary sarcoidosis is rare, and it isnecessary to distinguish its imaging features from lung cancer and tuberculosis. They may appear as an irregular mass with multiple nodules on thoracic computed tomography (CT). In this case, primary lung cancer was suspected in a 57-year-old papillary thyroid carcinoma patient, as the pulmonary lesions were non-radioiodine avid and progressed shortly afterward. The asymmetrical focal uptake that was demonstrated in integrin receptor imaging (


The present study investigated the intimate partner femicide (IPF) and intimate partner femicide–suicide (IPFS) perpetrators’ individual, relational, and contextual characteristics by analyzing, within the ecological approach, femicide cases that occurred in Italy from 2010 to 2019. On the topic, to date, scant studies examined possible differences between IPF and IPFS risk factors, and no studies have analyzed these factors by adopting an ecological systems model perspective. To this aim, archival research was carried out. Of a total of 1.207 femicides, 409 were IPF, and 227 were IPFS. Perpetrators’ age, level of employment, law enforcement membership, mental and/or physical illnesses, use of psychoactive substances, previous crimes, previous violent relationships, presence of children, previous violence in the couple, inability to accept the end of the relationship, quarrels and conflict, jealousy and the psychophysical illnesses of both authors and victims, as well as the use of firearms and victim’s request for help were analyzed. The results underlined the existence of different risk factors contributing to the IPF perpetrators’ decision to commit suicide such as perpetrators’ age, law enforcement membership, and firearm availability. These findings stress the need for specific risk assessment and management strategies for IPFS perpetrators. Considered the misogynous killing of women by men, motivated by hatred, contempt, pleasure, or a sense of ownership of women, femicide is the most extreme form of gender-based violence leading premature death of women  and is iIntimate partner femicide (IPF) is a pervasive form of violence against women and represents a serious social, criminal justice, and public health issue. It could be defined as “the killings of a woman by her intimate male partner, including current or former legal spouses, common-law partners, or boyfriends”  (p. 78).Previous empirical results highlighted that femicide committed by an intimate current or ex-partner in some cases, is followed by the perpetrator’s suicide ,7,8,9,10Unfortunately, while in recent years data on IPF have increased and have been widely acknowledged ,16,17, dPrevious studies on IPFS perpetrators’ risk factors showed that it is almost always characterized by an intimate male perpetrator and a female victim ,35,36,37At the individual level, existing studies focused on gender, age, race, alcohol/drug use, mental illness, and previous crimes committed by the author ,36,37,41Regarding IPFS perpetrators’ alcohol and drug use, instead, there are contrasting results ,30,36. AContrasting results also emerged regarding the presence or absence of the perpetrators’ illness, only sometimes reported in research ,12,21,41At the relational level, one of the most important predictors of IPFS was prior episodes of IPV: 72.0% were preceded by previous forced sexual intercourse, stalking, strangulation, and abuse during pregnancy . DespiteRegarding age, a study carried out in Ghana on 35 cases of IPFS , found tConcerning the presence of children, having one or more children in the household, who were not fathered by the perpetrator, was more common in IPFS cases , while aAt the contextual level, the most considered factor in IPFS was firearm availability from the perpetrator, ranging across different studies from 47.0% to 92.0% ,40,44,55Considering the devastating effects of these phenomena , to bettThe ecological approach is the most widely used model for understanding IPV and the factors associated with variations in its prevalence, e.g., ,62,63, a(1)Do male IPFS perpetrators, compared with only IPF perpetrators, differ from each other in terms of individual, relational, and contextual risk factors?(2)Are some individual, relational, and contextual risk factors predictive of the perpetrators’ suicide after IPF?In particular, we try to answer the following research questions:The current study was part of broader research on femicides called FATHERS , approved by the Departmental Ethics Committee (protocol nr. 18/2016).For the study, we conducted archival research by consulting online newspapers and collecting all femicide cases in Italy, from January 2010 to December 2019 following the Vienna Declaration definition . The anaWe considered IPFS in each case of IPF followed by the perpetrators’ suicide regardless of the amount of time elapsed between IPF and the suicide. In line with other studies and methodologies, information about IPF and IPFS cases and their characteristics were investigated by consulting databases of local, regional, and national online media and newspapers; those were the most suitable sources for collecting our data of interest and were able to provide us with as much information as we needed on these phenomena ,6,65, es. A total of three thousand ninety-two online articles were conferred using keywords . Furthermore, the authors monitored each IPF/IPFS case over time to detect any possible missing and additional information. From 1.207 femicides in Italy in the last decade, 636 were included and considered for further analyses as identified as IPF or IPFS cases.During data collection, every effort was made to record additional information about victims and perpetrators’ individual, relational, and contextual risk factors, the homicide and/or the subsequent suicide, and consulting NGO reports and the 27ora website Geographical data .Characteristics of the victims .Characteristics of the author .Characteristics related to the author/victim relationship .Criminological characteristics .For the present study, the following set of variables was analyzed:The perpetrators’ individual-level risk factors included socio-demographic information such as gender, age, level of employment , law enforcement membership, mental and physical illness, alcohol/drug use, previous crimes, and violent relationships.Among the relational risk factors that concern both the perpetrators and the victims were included: perpetrator-victim age gap, presence/absence of sons and/or daughters both in common and not in common, previous violence in the couple, inability to accept the end of the relationship, quarrels and conflict, jealousy and mental and/or physical illness both of perpetrators and victims.Finally, the contextual-level risk factors included the previous request for help from the victims and the use of a firearm as a weapon to commit IPF/IPFS.Data have been collected and codified according to a Fatality Review Protocol , developp < 0.05) was corrected according to the Bonferroni procedure in Step 1, Step 2, and Step 3.The data collected within the database were analyzed using the SPSS statistical package . Descriptive statistics were carried out to assess IPF/IPFS prevalence and characteristics. Then, we compared individual, relational, and contextual risk factors to investigate their possible role in some perpetrators’ decision to commit suicide after IPF by using odds ratios (ORs) because they are not influenced by sample size  ,68. FurtOf a total of 1.207 femicides that occurred in Italy in the last decade, 707 were IPF. A total of 636 cases were included in our sample, as no information about the perpetrators’ suicide intentions was found for the remaining 71 cases see .The perpetrator’s suicide followed 35.7% of IPF cases. In 88.5% of cases, perpetrators committed suicide within 24 h after the IPF. In contrast, in the remaining part of IPFS, perpetrators committed suicide in a time-lapse ranging from one day to seven months after IPF.Perpetrators’ age ranged from 18 to 97 years , while victims were aged between 16 and 93 years , 75.8% were older than their victims. On average male offenders were 7.04 (SD = 5.49) years older than their victims.Concerning IPF victims’ nationality, 77.4% were Italian. Regarding their occupational level, 27.2% resulted unemployed or retired, 27.0% had a low specialization level , and 4.2% had a high specialization level. A total of 9.3% of victims of IPF were affected by mental and/or physical illness.A total of 35.2% of perpetrators were unemployed or retired, 50.0% had low specialization occupations , and 9.7% were law enforcement officers. A total of 81.4% of perpetrators were Italian, 17.6% committed previous crimes, 12.6% used psychotropic substances such as alcohol, cocaine, or marijuana, and 8.8% of perpetrators were affected by a mental and/or physical illness. In 1.6% of IPF/IPFS cases, both the perpetrator and the victim suffered from a mental and/or physical illness.Regarding the characteristics of the relationship, in 84.3% of IPF cases, the perpetrator was the victim’s husband or ex-husband; in 57.7% of IPF cases, the perpetrator had a child in common with the victim, while in 21.5% of the cases, the children were only the victims. The most common motives for IPF were the perpetrator’s inability to accept the end of the relationship (35.7%) and jealousy (19.5%).A total of 38.5% of perpetrators had already been violent against the victim, and 29.6% used a firearm to commit IPF.We examined the possible existence of significant differences in terms of individual, relational, and contextual risk factors across IPF and IPFS perpetrators see .p < 0.001]. Respectively, 9.7% and 6.2% of suicide perpetrators used substances and committed previous crimes. However, perpetrators’ prior use of substances and/or involvement in criminal behaviors seem to be protective factors for suicide. Additionally, the history of violent relationships was a predictive risk factor only for IPF.At the individual level, the results showed that, respectively, 43.3% of low-specialized workers, 39.0% of highly specialized professionals, and 16.7% of law enforcement committed suicide after IPF. Only being part of law enforcement was a significant predictive factor for suicide after IPF . Among motives for IPF, the perpetrator’s and/or the victim’s mental and/or physical illness significantly increased the perpetrators’ likelihood of suicide .At the relational level, 64.8% of perpetrators with sons/daughters in common with the victim committed suicide after killing the women. To have sons/daughters in common was found to be a significant risk factor for perpetrators’ suicide after homicide .Concerning the contextual level, 55.5% of perpetrators used a weapon as a firearm for IPFS, compared to 15.2% for IPF. The use of a gun emerged as a significant predictive risk factor IPFS [OR = 6.98, 95% CI (4.79–10.17), A hierarchical logistic regression analysis was conducted to establish the predictive role of some individual, relational, and contextual risk factors in the IPF perpetrators’ decision to commit suicide.p < 0.01), being a member of law enforcement , and with suffering mental and/or physical illness . Interestingly, perpetrators suicide was negatively associated to substances use , to previous involvement in criminal behaviors  and to previous violent intimate relationships . At the relational level, perpetrators’ suicide emerged to be significantly associated to being older than the victims , having children in common , having children not in common  and with the presence of perpetrator and/or victim mental and/or physical illness (r = 0.20. p < 0.01). On the contrary, perpetrators’ jealousy  and quarrels and conflicts  were negatively associated to suicide after IPF. Regarding the contextual level risk factors, only using a firearm as a weapon to commit IPF was significantly associated with suicide .First, simple correlations were calculated between different risk factors and perpetrators’ suicide after IPF to analyze the relationship between variables. Regarding individual-level risk factors, the perpetrators’ suicide after IPF was found to be significantly associated with perpetrators’ age  and law enforcement membership were statistically significant individual risk factors for perpetrators’ suicide after IPF. In the second step, relational level risk factors were entered, significantly increasing the variance explained, through only quarrels and conflicts and jealousy were negatively associated with the IPF perpetrators’ suicide. In the third step, using a firearm to commit IPF was entered, indicating that holding and using a gun to kill the victim, being older, and being a law enforcement member were significant risk factors that predict IPFS. Additionally, quarrels/conflicts and jealousy were significantly and negatively associated with the IPF perpetrators’ suicide. The full model explained 26.0% of the total variance of the IPF perpetrators’ suicide.Intimate partner femicide is a widespread phenomenon worldwide , represeIn some cases, it could be followed by the perpetrator’s suicide ,32. HoweTo our knowledge, no studies analyzed and compared the IPF and IPFS perpetrators’ risk factors according to the ecological system theory ,60,61. STo this aim, we carried out an archival study on IPF in Italy in the last decade, of 636 cases of IPF, 227 were IPFS (35.7%).Following our first research question, the comparison between IPF and IPFS perpetrators’ risk factors highlighted, in line with prior studies, that using illicit drugs ,44,48,49Consistent with other studies on IPFS, the existence of a significant age gap between the perpetrator and the victim (7.04 years more for the perpetrator on average) ,12,37 anLikewise, as underlined by a previous study , no prioAmong motives for both IPF and IPFS, jealousy, quarrels and conflicts, and incapability to accept the end of the relationship were quite spread (even if they have not been found as risk factors for themselves) ,35,37,52Furthermore, it would seem that only the presence of the perpetrator’ and/or the victim’s psychophysical illnesses ,37,41,42About the second research question, the hierarchical logistic regression model results confirmed the perpetrators’ age role ,6,12 andThe present study has certain limitations. As in most archival research, information about IPF and IPFS was collected by searching and reading online newspaper articles, causing a high rate of missing cases as in the sample considered in the current study. Despite this, archival research using online newspaper articles is one of the methods adopted in this field ,6,65, esCausing the death of approximately two individuals, IPF and IPFS represent the most lethal forms of intimate partner abuse, leading to far-reaching effects on public health, families, friends, neighborhoods, and entire communities. This study contributes to the scant literature on the differences between IPF and IPFS fatal events and the need for prevention and intervention efforts. Consistent with other studies ,12, our Our results summarize the suicidal killer profile as an ordinary man without a history of criminal records or violent relationships, nor a previous history of drug or alcohol use . HoweverIf on one side, our results evidenced a suicidal killer as an ordinary man, and on the other side, they seem to suggest the need to better consider and investigate, consistently with Dawson , the posOur findings underline that law enforcement members and those who possess a firearm are more likely to commit suicide after the IPF.Regarding IPFS prevention, it could be crucial to focus on the employees’ psychological health, especially those equipped with a firearm. In fact, despite few studies, law enforcement members showed the presence of some job-related risk factors  that neeConsidering this, it could be helpful to develop, implement and evaluate the effectiveness of Behavioral Health Training functional to intercept signs, symptoms, and risk factors of mental health issues . BecauseFurthermore, further studies are needed considering the scant literature on the topic and the few variables considered to explain the phenomenon. Future studies should identify the critical risk factors related to the IPF perpetrator’s suicide and investigate which individual, relational and contextual dimensions operate as protective factors for the IPF perpetrators, by adopting quantitative and qualitative methods.For example, future research could focus on the contributing role of some cognitive and emotional dimensions associated with IPF perpetrators’ choice to commit suicide. IPFS perpetrators should have more to lose and be characterized by high levels of apprehension, fear of social disapproval, social margination, stigmatization of the consequences, and remorse related to killing the victim ,9,72,73.Finally, future research should be designed to collect multiple data sources , for exa


Graphene has unique properties that can be exploited for radiofrequency applications. Its characterization is key for the development of new graphene devices, circuits, and systems. Due to the two-dimensional nature of graphene, there are challenges in the methodology to extract relevant characteristics that are necessary for device design. In this work, the Thru-Reflect-Line (TRL) calibration was evaluated as a solution to extract graphene’s electrical characteristics from 1 GHz to 65 GHz, where the calibration structures’ requirements were analyzed. It was demonstrated that thick metallic contacts, a low-loss substrate, and a short and thin contact are necessary to characterize graphene. Furthermore, since graphene’s properties are dependent on the polarization voltage applied, a backgate has to be included so that graphene can be characterized for different chemical potentials. Such characterization is mandatory for the design of graphene RF electronics and can be used to extract characteristics such as graphene’s resistance, quantum capacitance, and kinetic inductance. Finally, the proposed structure was characterized, and graphene’s resistance and quantum capacitance were extracted. Two-dimensional 2D) materials have attracted researchers due to their unique properties. Their optical, electronic, and mechanical properties indicate that these materials have great potential for novel applications in electronics and optoelectronics ,2,3,4. AD materiaTo perform RF characterization, the first mandatory step consists of the vector network analyzer (VNA) calibration to shift the measurement plane to the device under test (DUT). That can be achieved through several available alternatives such as short-open-load-through (SOLT), line-reflect-match (LRM), and TRL . The TRLIn this work, the issues of implementing the TRL calibration applied to the characterization of 2D materials, such as graphene, is presented. The impact on the RF parameter extraction due to the design of the calibration structures, such as contact thickness, substrate loss, contacting graphene, and backgate design, are analyzed. Finally, resistance and quantum capacitance are extracted for graphene and processed using standard commercial 8-inch wafers processing equipment.This section discusses the implementation of the TRL calibration. It focuses on the design of the calibration structures, their frequency span, and the characteristics of the substrate. In this case, coplanar waveguides were used so that it is possible to contact them with ground signal ground RF probes and fabricate the device in small dimensions so that it can be used with graphene. Then, the determination of the characteristic impedance is discussed since the calibration is related to the impedance of the transmission lines. Finally, the extraction of the parameters used in this work is presented.To explore this calibration, it was necessary to define the calibration structures as well as the device under test. The number of lines used depends on the frequency span for which the calibration is used. The optimal line length is ¼ wavelength, and each thru/line pair has a usable bandwidth of 8:1 (frequency span/start frequency) ,19. Sinc3 Ω.cm, and the silicon oxide had a dielectric constant of 3.9 and loss tangent of 0.01. The metal used was copper, with a conductivity of 5.8 × 107 S/m. The dimensions , forward directivity (FD), and forward source match (FSM). It is now possible to obtain the S-parameter matrix [Three values obtained from the scikit-RF package  are usedr matrix :(4)SCPW=It is now possible to extract the characteristic impedance of the lines by transforming the S-parameters to T-parameters and through Equations (5) and (6) :(5)G=T12This method, however, is only accurate at lower frequencies. To obtain a more accurate approximation, the propagation constant that is given by the TRL calibration can be used by solving the capacitance and substrate conductance per unit length:The capacitance and conductance values are obtained from the lower frequency results of Equations (7) and (8), respectively, given that the method is more accurate in this range . Since tThe goal is to extract the series resistance, quantum capacitance, and kinetic inductance of graphene, since that information is necessary to develop interconnects, components, devices, and systems. Depending on the device geometry and number of graphene layers used, it is expected that the graphene device can have a capacitive or inductive behavior. To perform the TRL calibration, the NISTMultilineTRL procedure from the scikit-RF package  was usedThe proposed calibration method is carried out and tested in this section. Then, it is used to demonstrate different factors that can negatively impact or even prevent accurate measurements from being obtained. These are the metal thickness of the calibration structures and the substrate loss. Then, a calibration with thick copper contacts and high resistivity substrate is demonstrated, as well as including thin gold contacts and a buried gate for the characterization of graphene. Since the inclusion of the gold contacts and the buried gate are a modification of the original TRL calibration, their influence on the characteristic extraction is reported.The procedure starts with a typical calibration (such as SOLT) of the measurement system to shift the measurement planes to the transmission lines of the TRL calibration. Then, the TRL calibration structures are measured, and their characteristic impedance is extracted through the steps described in the previous section. In this case, the simulation software is used to simulate only the calibration structures, mimicking the initial step (SOLT and structure measurement for characteristic impedance extraction). After obtaining the characteristic impedance of the transmission line, the calibration can be carried out without any pre-calibration of the VNA, and the desired RF characteristics can be extracted. This corresponds to including two 2 m long, 53 Ω coaxial cables in the simulation, one for each measurement port. This demonstrates that the calibration can be performed even with slightly mismatched coaxial cables.Implementing this calibration with 2D materials can be a challenging task due to their physical dimensions and characteristics. Another constraint related to characterizing two-dimensional materials is their inherent high series resistance. Graphene has a sheet resistance of 0.5 kΩ to 1.5 kΩ . Since tAs described in , when grAn example of a graphene contact would be using a 20 nm thick Au contact, given that it allows for a small thickness to prevent graphene from rupturing and allows for a good contact resistance. In this case, to extract the RF characteristics, the TRL calibration method is evaluated. To test the effect of such small thickness on the calibration accuracy, the calibration was performed with 20 nm of Au. An accurate calibration would yield results close to 1 NH for the imaginary part and 1 kΩ for the real part. However, it is possible to observe that the inductance changes from approximately 0.2 nH to 0.7 nH, and the resistance is around 600 Ω. Therefore, it is possible to conclude that very thin contacts compromise the implementation of the TRL calibration and extraction of the device’s properties.Since the characteristics of the transmission line can influence the accuracy of the calibration procedure, the substrate conductivity can be another inaccuracy source. To demonstrate the impact of the substrate conductivity, a standard clean room doped silicon wafer with 100 Ω.cm is used. The measurement structures are as previously described, and the contacts are made of 5 µm thick copper to remove the previously described effect of the metallic resistance of the calibration accuracy. It is possible to observe that the substrate loss impacts the extraction of the RF characteristics of the device. The series resistance is in the 800 Ω to 900 Ω range when it should be around 1 kΩ, and the inductance is in the −165 nH to 0.8 nH range when 1 nH is expected. This is due to the increased transmission line losses that are induced by the substrate conductivity.As demonstrated, using thin metallic contacts and lossy substrate cause the calibration to provide very unreliable results. This is due to the extremely high losses that are introduced in the transmission line. To overcome these issues, two approaches must be taken: reduce the contact resistance and reduce the substrate conductivity. To reduce the contact resistance, copper was used, which has one of the highest conductivities, and its thickness was increased to 5 µm to further improve the contact. For the case of substrate conductivity, a commonly available 10 kΩ.cm silicon substrate was used.The problem of using 5 µm thick contacts is that these will break graphene, and contact will not be made. Therefore, a 20 nm gold contact with a length of 35 µm is included to bridge the connection from the 2D material to the copper contact. 7 S/m was included. These structures were simulated, and the characteristics of the lumped model were extracted. It is possible to observe that the accuracy of the measurements has increased when compared with the scenario where the structures have thin contacts, and the scenario where the substrate has a high conductivity. The extracted inductance is now around 0.9 nH to 1.1 nH, and the resistance is between 900 Ω to 1 kΩ, when 1 nH and 1 kΩ were expected, respectively. For example, when comparing with the high conductivity substrate scenario, where the inductance was much lower than 0.6 nH for frequencies lower than 25 GHz and the resistance was between 800 Ω to 900 Ω, this demonstrates that using 5 µm thick copper contacts and 10 kΩ·cm silicon substrate provides a reliable calibration for measuring 2D materials. Furthermore, 20 nm gold contact with the length of 35 µm and with a conductivity of 4.1 × 10Most applications of graphene use its conductivity that depends on its chemical potential. Such property can be modified through a voltage gating and has been explored for tunable applications . Therefo5 S/m and a thickness of 20 nm, resulting in 500 Ω/square. The DUT area corresponds to two squares in series, which totals 1 kΩ series resistance. To determine the characteristics of the mimic device, the TRL calibration without the backgate was performed as well as with a backgate made of gold. The previous simulations used a lumped element to mimic the extraction of the properties of graphene. That approach, however, does not account for the signal propagation in the device as well as its interaction with the backgate contact. Therefore, to mimic graphene, the lumped element was replaced by the best possible approach allowed by the simulator—a material with 1 × 104 S/m. An alternative could be implementing a backgate design that has a high impedance, which would prevent the RF signal propagation from being influenced by the backgate. However, this design would need to be fit for a frequency range from 1 GHz to 65 GHz. Since the purpose of the backgate is transmitting a DC voltage to gate the 2D material, a simpler solution would be to increase its resistance. The only constraint is that the gate oxide must be much more resistive than the series resistance of the backgate itself, so that the applied voltage at the gate is close to the applied voltage at the gate contact. Therefore, the same backgate design can be used if the material’s conductivity is modified. Since the highly conductive backgate contact impacts the parameter extraction, by reducing the backgate conductivity, it is possible to circumvent this problem. The extraction structures, with the same backgate design were simulated with a material with a conductivity of 2 × 10The proposed measurement and calibration structures were fabricated. In this case, the 2D material was graphene, and the device fabricated is shown in The fabricated structures were measured with the Keysight E5071C VNA , the GSG |Z|-Probes , and the CS-5 calibration substrate . Then, the previously proposed calibration procedure was undertaken, allowing us to extract graphene’s characteristics. These measurements were performed for 0 V, 5 V, 10 V, and 15 V of backgate bias. The extracted resistance and quantum capacitance of graphene are shown in 2V−1s−1, and the Dirac point is in the range of 6 to 8 V when it is gated with 15 nm of Al2O3, meaning that in this case, it should be from 50 to 67 V.The extracted resistance changes from around 450 Ω at 1 GHz down to around 250 Ω at 8.5 GHz for the bias voltage of 0 V. It then increases with the increase of the backgate voltage up to a maximum resistance of around 750 Ω at 1 GHz to around 400 Ω at 8.5 GHz, obtained at 15 V. The quantum capacitance changes from close to 0.55 pF at 1 GHz down to around 0.2 pF at 8.5 GHz for the bias voltage of 0 V. This capacitance reduces with the increase in voltage, achieving a capacitance of around 0.23 pF at 1 GHz down to around 0.1 pF at 8.5 GHz for the bias voltage of 0 V. Both resistive and capacitive behavior are as reported in the literature ,27. The Graphene is a promising material for several fields such as electronic and RF applications. The TRL calibration provides one of the highest calibration accuracies. In this work, the application of the TRL calibration for the extraction of the RF characteristics of graphene was explored. The implementation of the calibration was described, as well as the parameters that negatively affect its accuracy such as thin contacts and lossy substrate. To obtain an accurate calibration, thick contacts and low loss substrate should be used to reduce the losses in the transmission line. In this case, considering the characteristics of the transmission line, the series resistance is around 0.8 Ω, the substrate conductivity 10 kΩ·cm, and the backgate contact around 730 kΩ, which can be used as a reference point for the calibration. Since thick contacts will damage graphene, a short length thin contact was added, and its impact on the extraction of the RF characteristics of graphene was demonstrated to be negligible. Furthermore, since graphene’s conductivity changes according to its chemical potential, a backgate was added to characterize it for different potentials. Finally, the proposed structures were fabricated and measured, and the properties of graphene were extracted, from 1 GHz to 8.5 GHz. Its resistance at 1 GHz changed from around 450 Ω to 750 Ω at 0 V and 15 V, respectively; at 8.5 GHz, it changed from around 250 Ω to 400 Ω at 0 V and 15 V, respectively. Its quantum capacitance at 1 GHz changed from around 0.55 pF to 0.23 pF at 0 V and 15 V, respectively; at 8.5 GHz, it changed from around 0.2 pF to 0.1 pF at 0 V and 15 V, respectively. The development of such calibration procedure to characterize graphene will allow us to further improve the knowledge of the graphene’s RF characteristics, paving the way to obtain tunable electronics either through biasing or biological functionalization. Finally, the proposed calibration and extraction method are not limited to graphene and can be used with other 2D materials.


The persistence of inflammatory mediators in tissue niches significantly impacts regenerative outcomes and contributes to chronic diseases. Interleukin-4 (IL4) boosts pro-healing phenotypes in macrophages (Mφ) and triggers the activation of signal transducer and activator of transcription 6 (STAT6). Since the IL4/STAT6 pathway reduces Mφ responsiveness to inflammation in a targeted and precise manner, IL4 delivery offers personalized possibilities to overcome inflammatory events. Despite its therapeutic potential, the limited success of IL4-targeted delivery is hampered by inefficient vehicles. Magnetically assisted technologies offer precise and tunable nanodevices for the delivery of cytokines by combining contactless modulation, high tissue penetration, imaging features, and low interference with the biological environment. Although superparamagnetic iron oxide nanoparticles (SPION) have shown clinical applicability in imaging, SPION-based approaches have rarely been explored for targeted delivery and cell programming. Herein, we hypothesized that SPION-based carriers assist in efficient IL4 delivery to Mφ, favoring a pro-regenerative phenotype (M2φ). Our results confirmed the efficiency of SPION-IL4 and Mφ responsiveness to SPION-IL4 with evidence of STAT6-mediated polarization. SPION-IL4-treated Mφ showed increased expression of M2φ associated-mediators  when compared to the well-established soluble IL4. The ability of SPION-IL4 to direct Mφ polarization using sophisticated magnetic nanotools is valuable for resolving inflammation and assisting innovative strategies for chronic inflammatory conditions. Inflammation is a dynamic process triggered by the immune system in response to injury or infection. However, the uncontrolled activation of immune cells results in persistent inflammatory signals, with deleterious consequences for tissue regeneration. Resolving inflammation is thus a challenge to be overcome with impact in the management of chronic inflammatory diseases , autoimmInterleukin-4 (IL4) is a key regulator in humoral and adaptive immunity and a well-known promoter of alternatively activated M2 macrophages (M2φ) . The preMagnetically assisted technologies ,14,15 ofAlthough IL4 is a well-established M2φ switch, there is a lack of research on IL4 delivery using particulate carriers. Such systems would enable cell and receptor magnetic targeting strategies for the in situ modulation of the macrophage phenotype to locally discourage abnormal inflammatory signals and reestablish pro-healing environments. Therefore, we hypothesized that superparamagnetic iron oxide nanoparticles (SPION) decorated with IL4 (SPION-IL4) favor M2φ via magnetically guided IL4 presentation. By targeting Mφ, which has a coordinated action over inflammatory cascades, SPION-IL4 could assist in the contactless control of inflammatory cues in injured or pathological environments. Here, SPION-IL4 were fabricated and investigated downstream of the IL4 receptor in Mφ. The activation of STAT6 and the gene and protein inflammatory mediators were assessed with different concentrations of SPION-IL4 and compared with carrier-free IL4 (Exo IL4) exogenously supplemented to the culture medium. We foresee improved and more effective SPION-mediated therapeutics in precision Mφ targeting for the immunomodulation of persistent inflammatory environments.w/w and that of the organic component was 22 ± 9% w/w, confirming the presence of dextran-COOH in the shell of the SPION  carbodiimide/N-hydroxysuccinimide (EDC/NHS) chemistry. The magnetic response of SPION-IL4 to EMF was evaluated, and SPION-IL4 exhibited magnetic responsiveness A. The inhe SPION .A stable amide (-CONH-) covalent linkage was selected for the magnetic presentation of IL4 to macrophages. This conjugation method has been used for the functionalization of nanomaterials with proteins, peptides, antibodies, and nucleic acids ,17,18,19−1, which is due to the stretching vibrations of the carbonyl bond of the carboxylic acid group (COOH) in dextran. When comparing the SPION to the SPION-IL4 spectrum, this band was replaced by two bands at ~1650 cm−1 and ~1550 cm−1. The band at ~1650 cm−1 is due to carbonyl bond stretching vibrations, whereas the band at 1550 cm−1 represents N–H bond bending vibrations. Thus, the substitution of a COOH representative band in SPION by two amide bond representative bands indicates the successful functionalization of SPION with IL4. The absence of a 15 kDa band associated with IL4 molecular weight in the sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE)  spectroscopy D. The FTDS-PAGE)  in the SDS-PAGE) E. DespitDS-PAGE) .2 [p > 0.05) were observed in cellular viability  administration to Mφ. Thus, we estimated a 646 fg IL4/cell in SPION-IL4/30, and a 1940 fg IL4/cell in both SPION-IL4/100 and Exo IL4 conditions. Hence, SPION-IL4/100 corresponded to the highest SPION concentration to be administered to cells in the present study, with an equivalent number of IL4 molecules to Exo IL4, namely 1940 fg IL4/cell.Unidirectional formation of highly oriented rods of SPION-IL4 was observed in the SEM images A, suggesp < 0.0001). When SPION-IL4 or soluble IL4 binds to the IL4 receptor, the receptor is no longer available for immunostaining. Thus, the signal detected was related to IL4 receptors that were not previously occupied by IL4. Since SPION-IL4/30 is related to a lower concentration of both SPION and IL4 in comparison to SPION-IL4/100 and Exo IL4, more free receptors are available, and thus the signal intensity is higher. Despite the successful delivery of IL4 via SPION-IL4 to the IL4 cognate receptor, the concentrations of SPION-IL4 used did not saturate all the IL4-receptors available in the cells.To investigate the SPION-IL4 formulation, interactions with IL4 receptor cells were immunostained with an antibody against IL4 receptor alpha (IL4Rα) B. SPION-p < 0.05 and p < 0.01) (p > 0.05) independently of the iron concentration used  , and ind < 0.01) D-II. Theion used D-II. Conion used E.Overall, SPION-IL4 was more effective in the transport and delivery of IL4 participating in the activation of the IL4/STAT6 pathway. The production of IL4 at the gene and post-transcript levels was more efficient when magnetic nano-vehicles were applied, especially with a lower concentration of SPION-IL4 (30 µg/mL), suggesting that higher concentrations of IL4, either provided by soluble IL4 (Exo IL4) or in SPION-IL4 nanocarriers, may lead to suboptimal levels of IL4 mediated signals in THP1-derived macrophages.+ cells was observed after 24 h in SPION-IL4 conditions. The decreased availability of the CD11b receptor on cells treated with SPION-IL4 was accompanied by an elongated morphology (white arrow), which has been associated with a M2φ phenotype [According to the literature, integrin αM (CD11b) has been implicated in the activation of pro-inflammatory transcription factors, such as nuclear factor kappa B (NFKB) , as wellhenotype .The cocktail of cytokines/chemokines released by macrophages is indispensable for characterizing their polarized states. The cytokine/chemokine profile of SPION-IL4 treated cells was reported for pro- and anti-inflammatory mediators B. IL8 isThe concentration of SPION-IL4 seemed to have a particular effect on the secreted chemokines CCL5 (C-C Motif Chemokine Ligand-5) and CCL2 (C-C Motif Chemokine Ligand-2). The production of CCL5, whose signaling contributes to M1φ polarization and can be inhibited by IL4 , was detp < 0.05) also supports an M2φ shift.Since M2φ polarization is associated with decreased inducible nitric oxide synthase (iNOS) expression and increased ARG1 levels , the ratIn this study, we investigated the promising role of magnetic nanoparticle-mediated delivery of IL4 aiming at M2φ polarization. SPION-IL4 contributed to IL4-mediated actions and to the expression of STAT6 responsive molecules such as CCL2 and ARG1. In particular, ARG1 together with STAT6 has been described to modulate Mφ phenotypes and to accelerate inflammation resolution . Studies3O4) and a dextran -COOH shell were purchased from micromod GmbH . SPION were functionalized with IL4 (SPION-IL4) by carbodiimide coupling chemistry using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC)  and N-hydroxysuccinimide (NHS) . All production steps were performed at room temperature (RT) unless stated otherwise. SPION were mixed with EDC and NHS in 0.5 M MES buffer  (pH 6.3) for 45 min. Then, activated SPION were washed by magnetic separation with 0.01 M phosphate buffered saline (PBS)  (pH = 7.4), and mixed with 10 µg of Recombinant Human IL4  for 3 h. Following a washing step, 25 mM glycine  in 0.01 M PBS was added, and the mixture was continuously stirred for 30 min. SPION-IL4 was washed three times and resuspended in 1 mL of ultra-pure water. SPION-IL4 was further purified by dialysis  against water for 24 h and stored at 4 °C.SPION composed of a magnetite core  and after functionalization (SPION-IL4). An ultrapure water background was used before the experiments. SPION-IL4 morphology were analyzed using scanning transmission electron microscopy (STEM) . The elemental composition of SPION-IL4 was determined using Energy Dispersive X-ray spectroscopy (EDS) . The hydrodynamic diameter and zeta potential of SPION-IL4 were determined using a Malvern NanoZS .Fourier transform infrared (FTIR) spectra were recorded with an IRPrestige 21 spectrometer (Shimadzu) using the attenuated total reflectance (ATR) mode over the range of 4000–600 cm®20 detergent  and incubated with Anti-Rabbit IgG secondary antibody  at RT for 1 h. After washing in TBST, the signal was developed using an AP Conjugate Substrate Kit  according to the manufacturer’s instructions. For the western blotting assay, the supernatant of the SPION-IL4 solution (S) after brief centrifugation was also included as a control for IL4 binding to SPION. Membrane images were acquired using a digital scanner  in conjunction with Epson Scan software ver. 3.9.2.1US. IL4 bands were further quantified using ImageJ 1.50i  software.The functionalization efficiency of IL4 for SPION was evaluated using western blotting. Samples: SPION, SPION-IL4, supernatant from the IL4 incubation solution with SPION (S1), and supernatant from the first washing step with PBS (S2) were analyzed, as well as standard IL4 solutions (0 to 25 µg/mL). A mixture of Laemmli buffer  and the samples (1:1) was incubated at 95 °C for 5 min before running on a 12.5% separation gel and 4% stacking gel . The PageRuler Plus Prestained Protein Ladder  was used as a molecular weight marker. The gel was stained with Coomassie Brilliant Blue  for protein visualization. The gel was transferred to a nitrocellulose membrane  using a Pierce Power System . The membranes were incubated for 1 h with a blocking solution, followed by overnight incubation with recombinant anti-IL4  at 4 °C. Membranes were washed in Tris-Buffered Saline containing 0.1% Tween2 atmosphere. THP1-derived macrophages were differentiated with 100 nM phorbol 12-myristate 13-acetate  for 24 h, followed by 24 h incubation with a PMA-free medium. Adherent THP1-derived macrophages were cultured and expanded in RPMI medium. Two time-points (1 h or 24 h) and up to four conditions were investigated: THP1 cells cultured with (i) SPION-IL4 at 30 µg/mL or (ii) SPION-IL4 at 100 µg/mL of iron  and non-functionalized SPION at (iii) 30 µg/mL or (iv) 100 µg/mL of iron . Conditions (iii) and (iv) act as experimental controls for IL4 functionalization. Magnetically stimulated THP1 cells exogenously supplemented with soluble IL4 were used as a positive control (hereafter referred to as Exo IL4) whereas cells cultured on tissue culture polystyrene (TCPs) were used as a negative control. The concentration of IL4 per cell used in the SPION-IL4/100 and Exo IL4 conditions was the same. EMF was provided by a magnefect nano device  (350 mT/well) under all conditions.Human monocytes ) were cultured in Roswell Park Memorial Institute medium (RPMI)  supplemented with 1% L-Glutamine  in a humidified 5% CO× 105 cells per well , followed by incubation with SPION-IL4 at 37 °C under magnetic stimulation. For LDH experiments, cells were seeded at 1 × 104 cells per well . Live/dead cell double staining: Cells were washed with PBS and incubated with 2 µM calcein AM in PBS  and 4 µM propidium iodide (PI)  in the dark for 30 min at 37 °C. Cells were rinsed in PBS, and the fluorescence signal was acquired using a transmitted and reflected light microscope with Apotome 2 . Cell viability was assessed using both 3--5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) and LDH assays. MTS: THP1 cells were incubated in MTS solution  for 3 h at 37 °C in a 5% CO2 atmosphere and protected from light. The supernatant was transferred to a new 96-well plate and the optical absorbance was measured at 490 nm . LDH: At the respective time-points, cytotoxicity was measured in each sample medium using a CyQuant LDH Cytotoxicity Assay kit , following the manufacturer’s instructions. LDH activity was measured at 490 nm/680 nm excitation/emission wavelengths using a microplate reader .To determine the effect of SPION-IL4 on cells, THP1-derived macrophages were seeded at 1 ® RNA Isolation Reagent  following the manufacturer’s instructions. RNA was quantified using a Nanodrop® ND-1000 spectrophotometer  at 260/280 nm. First-strand complementary DNA was synthesized from 1 µg of RNA from each sample  in a 20 µL reaction using a Mastercycler® ep realplex gradient S machine (Eppendorf). Transcripts were quantified by quantitative polymerase chain reaction (qPCR) using the PerfeCTA SYBR Green FastMix kit  following the manufacturer’s protocol in a Real-Time Mastercycler ep realplex thermocycler (Eppendorf). Primer sequences  phosphorylation (pSTAT6) used to screen the effects of SPION-IL4 in THP1-derived macrophages was assessed using a cell-based phospho-STAT6 (pTyr641) ELISA kit  following the manufacturer’s instructions.IL4 Receptor Alpha (IL4Rα) is the alpha chain of the IL4 receptor and is involved in the regulation of alternative activation of macrophages and, consequently, IL4 secretion levels, whereas CD11b is an integrin family member involved in the regulation of leukocyte adhesion and migration to mediate the inflammatory response. To detect these molecules, THP1-derived macrophages were fixed in 4% neutral buffered formalin  overnight and stored in PBS at 4 °C until use. Subsequently, the cells were incubated with 0.025% Triton-X100  in PBS, and blocking was performed using 2.5% normal horse serum . Cells were incubated overnight at 4 °C with anti-IL4Rα , anti-IL4 , or anti-CD11b  diluted in 0.1% bovine serum albumin diluted , followed by incubation for 1 h at RT with the secondary antibody . The samples were washed three times with PBS before staining with DAPI  for 10 min. Immune detection was then assessed by confocal laser scanning microscopy, and 3D reconstruction images obtained from the z-stacks were obtained using TCS SP8 software . IL4Rα and IL4 fluorescence intensities were quantified using ImageJ 1.50i software .In the case of CD11b, the number of cells positive for CD11b was determined by flow cytometry. The cells were permeabilized and incubated with anti-CD11b  for 20 min, followed by incubation with the secondary antibody (Anti-Rabbit IgG Alexa Fluor 488) for 30 min in the dark. Cells were then washed with PBS and resuspended in acquisition buffer  before acquisition on a FACSAria III sorter equipped with blue and red lasers . The cells were identified using forward and side scattering. A minimum of 5000 cells were acquired and analyzed using the FACS Diva software version 7. Unstained cells were used to establish cell autofluorescence, and cells incubated only with secondary antibodies were used as controls. The population of cells positive for CD11b was expressed as a percentage.Scanning electron microscopy (SEM) was used to visualize the physical distribution of SPION-IL4 in the cells. THP1-derived macrophages were fixed in 4% neutral buffered formalin  overnight and dehydrated using ascending concentrations of ethanol (from 30 to 100%), followed by a 5 min immersion in hexamethyldisiloxane . The samples were then air-dried overnight and sputter-coated  with gold. Images were acquired using scanning electron microscopy .× 105 THP1-derived macrophages per well were cultured in 24 wells plates  before supplementing with SPION-IL4 and IL4. After 24 h, the supernatants were collected and analyzed using a Proteome Profiler™ Human Cytokine Array Kit  following the manufacturer’s instructions. The spot intensity was detected using the Odyssey Fc Imaging System 2800 (LI-COR®) and quantified using ImageJ 1.52o  software. Supernatants were also analyzed for the quantification of secreted cytokines using commercially available enzyme immunoassay kits for ARG1  and IL4  following the manufacturer’s instructions.For the cytokine arrays, 3 (n = 3), except for flow cytometry analysis of CD11b, which is representative of two independent experiments (n = 2), with a minimum of 5000 cells analyzed per experiment. Statistical analysis was performed using SPSS statistical software (version 27.0.1.0). First, a Shapiro–Wilk test was used to ascertain data normality and Levene’s test for homogeneity of variances. Normality and variance homogeneity were rejected, and non-parametric tests were used . Different degrees of confidence were considered, p < 0.05, p < 0.01, p < 0.0001, and are represented by symbols * and # for p < 0.05, **, ## for p < 0.01, and **** for p < 0.0001.Results are expressed as the mean ± standard error of the mean (SE), representative of three independent experiments This study showed that SPION-IL4 is a suitable IL4 presenter, anticipating the modulation of cell behavior with insights into macrophage-priming strategies mediated by EMF-actuated SPION. SPION-IL4 treated Mφ showed an increased expression of genes such as IL10 and ARG1 and proteins (CCL2 and IL1Ra) typically associated with the M2φ phenotype, with unique advantages over carrier-free administration. Together with the therapeutic effect, the magnetic memory of SPION-IL4 comprises multimodal possibilities, including cell tracking and in vivo identification of IL4-targeted populations.In view of the presence of macrophages in almost all tissues and organs, SPION in combination with IL4 and/or other cytokines could hold wide applicability as phenotype modulators and, consequently, as magnetically responsive switches of pro- and anti-inflammatory activities. Either in the form of magnetic delivery systems or magnetically responsive elements incorporated into tridimensional matrices, these systems could render immunomodulatory actions foreseeing improved therapeutic solutions for chronic inflammation conditions and tissue regeneration.