Publications

The interaction of nanoparticles with biological media is a topic of general interest for drug delivery systems and among those for active nanoparticles, also called nanomotors. Herein, we report the use of super resolution microscopy, in particular, stochastic optical reconstruction microscopy (STORM), to characterize the formation of a protein corona around active enzyme-powered nanomotors. First, we characterized the distribution and number of enzymes on nano-sized particles and characterized their motion capabilities. Then, we incubated the nanomotors with fluorescently labelled serum proteins. Interestingly, we observed a significant decrease of protein corona formation (20%) and different composition, which was studied by proteomic analysis. Moreover, motion was not hindered, as nanomotors displayed enhanced diffusion regardless of the protein corona. Elucidating how active particles interact with biological media and maintain their self-propulsion after protein corona formation will pave the way for the use of these systems in complex biological fluids in biomedicine.; The interaction of self-propelled nanomotors with biological media is of outmost relevance when considering their actuation within biological contexts. Here, we explored how protein corona forms around active nanomotors using STORM.

JTD Keywords: Gold, Impact, Nanoparticle uptake, Plasma, Size, Surface-properties

Although multifunctional wearable devices have been widely investigated for healthcare systems, augmented/virtual realities, and telemedicines, there are few reports on multiple signal monitoring and logical signal processing by using one single nanomaterial without additional algorithms or rigid application-specific integrated circuit chips. Here, multifunctional intelligent wearable devices are developed using monolithically patterned gold nanowires for both signal monitoring and processing. Gold bulk and hollow nanowires show distinctive electrical properties with high chemical stability and high stretchability. In accordance, the monolithically patterned gold nanowires can be used to fabricate the robust interfaces, programmable sensors, on-demand heating systems, and strain-gated logical circuits. The stretchable sensors show high sensitivity for strain and temperature changes on the skin. Furthermore, the micro-wrinkle structures of gold nanowires exhibit the negative gauge factor, which can be used for strain-gated logical circuits. Taken together, this multifunctional intelligent wearable device would be harnessed as a promising platform for futuristic electronic and biomedical applications.© 2023 Wiley-VCH GmbH.

JTD Keywords: electronics, fabrication, intelligent multifunction, monolithic patterns, signal monitoring and processing, wearable devices, Gold nanowires, Intelligent multifunction, Intraocular-pressure, Monolithic patterns, Signal monitoring and processing, Wearable devices

Alzheimer's disease is characterized by a combination of several neuropathological hallmarks, such as extracellular aggregates of beta amyloid (Aβ). Numerous alternatives have been studied for inhibiting Aβ aggregation but, at this time, there are no effective treatments available. Here, we developed the tri-component nanohybrid system AuNPs@POM@PEG based on gold nanoparticles (AuNPs) covered with polyoxometalates (POMs) and polyethylene glycol (PEG). In this work, AuNPs@POM@PEG demonstrated the inhibition of the formation of amyloid fibrils, showing a 75% decrease in Aβ aggregation in vitro. As it is a potential candidate for the treatment of Alzheimer's disease, we evaluated the cytotoxicity of AuNPs@POM@PEG and its ability to cross the blood-brain barrier (BBB). We achieved a stable nanosystem that is non-cytotoxic below 2.5 nM to human neurovascular cells. The brain permeability of AuNPs@POM@PEG was analyzed in an in vitro microphysiological model of the BBB (BBB-on-a-chip), containing 3D human neurovascular cell co-cultures and microfluidics. The results show that AuNPs@POM@PEG was able to cross the brain endothelial barrier in the chip and demonstrated that POM does not affect the barrier integrity, giving the green light to further studies into this system as a nanotherapeutic.

JTD Keywords: beta-amyloid, blood-brain barrier organ-on-a-chip, cellular uptake, citrate, cytotoxicity, electrocatalytic reduction, gold nanoparticles, hypothesis, nanorods, polyoxometalates, size, stability, surface, Alzheimers-disease, Blood–brain barrier organ-on-a-chip, Gold nanoparticles, Nanovehicle, Polyoxometalates, Β-amyloid

This work presents a novel nanoparticle-based thermosensor implant able to reveal the precise temperature variations along the polymer filaments, as it contracts and expands due to changes in the macroscale local temperature. The multimodal device is able to trace the position and the temperature of a polypropylene mesh, employed in abdominal hernia repair, by combining plasmon resonance and Raman spectroscopy with hydrogel responsive system. The novelty relies on the attachment of the biocompatible nanoparticles, based on gold stabilized by a chitosan-shell, already charged with the Raman reporter (RaR) molecules, to the robust prosthesis, without the need of chemical linkers. The SERS enhanced effect observed is potentiated by the presence of a quite thick layer of the copolymer (poly(N-isopropylacrylamide)-co-poly(acrylamide)) hydrogel. At temperatures above the LCST of PNIPAAm-co-PAAm, the water molecules are expulsed and the hydrogel layer contracts, leaving the RaR molecules more accessible to the Raman source. In vitro studies with fibroblast cells reveal that the functionalized surgical mesh is biocompatible and no toxic substances are leached in the medium. The mesh sensor opens new frontiers to semi-invasive diagnosis and infection prevention in hernia repair by using SERS spectroscopy. It also offers new possibilities to the functionalization of other healthcare products.© 2023 Wiley-VCH GmbH.

JTD Keywords: adhesion, blends, chitosan, gold nanoparticles, poly(n-isopropylacrylamide), polypropylene mesh, polypropylene meshes, repair, scattering, silver, surgical implants, thermosensitive hydrogels, toxicity, Chitosan, Gold nanoparticles, Polypropylene meshes, Surgical implants, Thermosensitive hydrogels

The lack of predictive models that mimic the blood-brain barrier (BBB) hinders the development of effective drugs for neurodegenerative diseases. Animal models behave differently from humans, are expensive and have ethical constraints. Organ-on-a-chip (OoC) platforms offer several advantages to resembling physiological and pathological conditions in a versatile, reproducible, and animal-free manner. In addition, OoC give us the possibility to incorporate sensors to determine cell culture features such as trans-endothelial electrical resistance (TEER). Here, we developed a BBB-on-a-chip (BBB-oC) platform with a TEER measurement system in close distance to the barrier used for the first time for the evaluation of the permeability performance of targeted gold nanorods for theranostics of Alzheimer's disease. GNR-PEG-Ang2/D1 is a therapeutic nanosystem previously developed by us consisting of gold nanorods (GNR) functionalized with polyethylene glycol (PEG), angiopep-2 peptide (Ang2) to overcome the BBB and the D1 peptide as beta amyloid fibrillation inhibitor, finally obtaining GNR-PEG-Ang2/D1 which showed to be useful for disaggregation of the amyloid in in vitro and in vivo models. In this work, we evaluated its cytotoxicity, permeability, and some indications of its impact on the brain endothelium by employing an animal-free device based on neurovascular human cells.In this work, we fabricated a BBB-oC with human astrocytes, pericytes and endothelial cells and a TEER measuring system (TEER-BBB-oC) integrated at a micrometric distance of the endothelial barrier. The characterization displayed a neurovascular network and the expression of tight junctions in the endothelium. We produced GNR-PEG-Ang2/D1 and determined its non-cytotoxic range (0.05-0.4 nM) for plated cells included in the BBB-oC and confirmed its harmless effect at the highest concentration (0.4 nM) in the microfluidic device. The permeability assays revealed that GNR-PEG-Ang2/D1 cross the BBB and this entry is facilitated by Ang2 peptide. Parallel to the permeability analysis of GNR-PEG-Ang2/D1, an interesting behavior of the TJs expression was observed after its administration probably related to the ligands on the nanoparticle surface.BBB-oC with a novel TEER integrated setup which allow a correct read-out and cell imaging monitoring was proven as a functional and throughput platform to evaluate the brain permeability performance of nanotherapeutics in a physiological environment with human cells, putting forward a viable alternative to animal experimentation.© 2023. The Author(s).

JTD Keywords: alzheimer disease (ad), cell-culture, cytotoxicity, endothelial-cells, gold nanoparticles, microfluidic platform, model, organ-on-a-chip (ooc), peptide, tight junction, trans-endothelial electrical resistance (teer), transport, Alzheimer disease (ad), Blood-brain barrier (bbb), Blood-brain-barrier, Blood–brain barrier (bbb), Gold nanoparticles, Organ-on-a-chip (ooc), Trans-endothelial electrical resistance (teer)

The quest for novel nanoscale materials for different applications necessitates that they are easy to obtain and have excellent physical properties and low toxicity. Moreover, considering the ongoing environmental impact of noxious chemical waste products, it is important to adopt eco-friendly approaches for nanoparticle synthesis. In this work, a natural polymer (medium molecular weight chitosan) derived from chitin was employed as a reducing agent to obtain gold nanoparticles (AuNPs) with a chitosan shell (AuNPs@CS) by a microwave oven. The chitosan is economically viable and cost-competitive in the market showing also nontoxic behavior in the environment and living organisms. The synthesized AuNPs@CS-FITC NPs were fully characterized by spectroscopic and microscopic characterization techniques. The size distribution of NPs was about 15 nm, which is a suitable dimension to use in biomedical applications due to their high tissue penetration, great circulation in blood, and optimal clearance as well as low toxicity. The prepared polymer-capped NPs were further functionalized with a fluorescent molecule, i.e., Fluorescein-5-isothiocyanate (FITC), to perform imaging in the cell. The results highlighted the goodness of the synthesis procedure, as well as the high internalization rate that resulted in an optimal fluorescence intensity. Thus, this work presents a good sustainable/green approach-mediated polymer nanocomposite for various applications in the field of diagnostic imaging.

JTD Keywords: Agent, Biomedical, Fluorescent, Gold nanoparticles, Green synthesis, Nanomaterials, Silver nanoparticles

Polypropylene (PP) surgical meshes, with different knitted architectures, were chemically functionalized with gold nanoparticles (AuNPs) and 4-mercaptothiazole (4-MB) to transform their fibers into a surface enhanced Raman scattering (SERS) detectable plastic material. The application of a thin layer of poly[N-isopropylacrylamide-co-N,N'-methylene bis(acrylamide)] (PNIPAAm-co-MBA) graft copolymer, covalently polymerized to the mesh-gold substrate, caused the conversion of the inert plastic into a thermoresponsive material, resulting in the first PP implantable mesh with both SERS and temperature stimulus responses. AuNPs were homogeneously distributed over the PP yarns, offering a clear SERS recognition together with higher PNIPAAm lower critical solution temperature (LCST ∼ 37 °C) than without the metallic particles (LCST ∼ 32 °C). An infrared thermographic camera was used to observe the polymer-hydrogel folding-unfolding process and to identify the new value of the LCST, connected with the heat generation by plasmonic-resonance gold NPs. The development of SERS PP prosthesis will be relevant for the bioimaging and biomarker detection of the implant by using the plasmonic effect and Raman vibrational spectroscopy for minimally invasive interventions (such as laparoscopy), to prevent patient inflammatory processes. Furthermore, Raman sources have been proved to not damage the cells, like happens with near-infrared irradiation, representing another advantage of moving to SERS approaches. The findings reported here offer unprecedented application possibilities in the biomedical field by extrapolating the material functionalization to other nonabsorbable polymer made devices (e.g., surgical sutures, grapes, wound dressings, among others).

JTD Keywords: gold nanoparticles, poly(n-isopropylacrylamide), polymers, polypropylene, raman-spectroscopy, reduction, resonance, sers spectroscopy, size, surface functionalization, Gold nanoparticles, Polypropylene, Surface functionalization

The influence of the properties of different solid substrates on the tethering of two antibodies, IgG1-CR3022 and IgG1-S309, which were specifically engineered for the detection of SARS-CoV-2, has been examined at the molecular level using conventional and accelerated Molecular Dynamics (cMD and aMD, respectively). Two surfaces with very different properties and widely used in immunosensors for diagnosis, amorphous silica and the most stable facet of the face-centered cubic gold structure, have been considered. The effects of such surfaces on the structure and orientation of the immobilized antibodies have been determined by quantifying the tilt and hinge angles that describe the orientation and shape of the antibody, respectively, and the dihedrals that measure the relative position of the antibody arms with respect to the surface. Results show that the interactions with amorphous silica, which are mainly electrostatic due to the charged nature of the surface, help to preserve the orientation and structure of the antibodies, especially of the IgG1-CR3022, indicating that the primary sequence of those antibodies also plays some role. Instead, short-range van der Waals interactions with the inert gold surface cause a higher degree tilting and fraying of the antibodies with respect to amorphous silica. The interactions between the antibodies and the surface also affect the correlation among the different angles and dihedrals, which increases with their strength. Overall, results explain why amorphous silica substrates are frequently used to immobilize antibodies in immunosensors. © 2022 The Authors

JTD Keywords: amorphous silica, antibody immobilization, enzyme, gol d, gold, immobilization, immunosensor, molecu l a r dynamics, molecular dynamics, protein adsorption, sars-cov-2 immunosensor, simulations, spike protein, surface interactions, target, vaccine, Amorphous silica, Antibodies, Antibody engineering, Antibody immobilization, Antibody structure, Article, Chemical detection, Computer model, Controlled study, Dihedral angle, Gold, In-silico, Molecular dynamics, Molecular levels, Molecular-dynamics, Nonhuman, Property, Sars, Sars-cov-2 immunosensor, Severe acute respiratory syndrome coronavirus 2, Silica, Silico studies, Silicon dioxide, Solid substrates, Structure analysis, Substrate chemistry, Substrates, Van der waals forces, Virus detection

Charge separation and transport through the reaction center of photosystem I (PSI) is an essential part of the photosynthetic electron transport chain. A strategy is developed to immobilize and orient PSI complexes on gold electrodes allowing to probe the complex's electron acceptor side, the chlorophyll special pair P700. Electrochemical scanning tunneling microscopy (ECSTM) imaging and current-distance spectroscopy of single protein complex shows lateral size in agreement with its known dimensions, and a PSI apparent height that depends on the probe potential revealing a gating effect in protein conductance. In current-distance spectroscopy, it is observed that the distance-decay constant of the current between PSI and the ECSTM probe depends on the sample and probe electrode potentials. The longest charge exchange distance (lowest distance-decay constant ?) is observed at sample potential 0 mV/SSC (SSC: reference electrode silver/silver chloride) and probe potential 400 mV/SSC. These potentials correspond to hole injection into an electronic state that is available in the absence of illumination. It is proposed that a pair of tryptophan residues located at the interface between P700 and the solution and known to support the hydrophobic recognition of the PSI redox partner plastocyanin, may have an additional role as hole exchange mediator in charge transport through PSI.© 2021 Wiley-VCH GmbH.

JTD Keywords: azurin, current distance decay spectroscopy, cytochrome c(6), electrochemical scanning tunneling microscopy (ecstm), electrochemistry, photosystem i, photosystem-i, plastocyanin, protein electron transfer, recognition, single metalloprotein, single molecules, structural basis, tunneling spectroscopy, 'current, Amino acids, Charge transfer, Chlorine compounds, Current distance decay spectroscopy, Decay spectroscopies, Distance decay, Electrochemical scanning tunneling microscopy, Electrochemical scanning tunneling microscopy (ecstm), Electrodes, Electron transfer, Electron transport properties, Gold compounds, Photosystem i, Photosystems, Protein electron transfer, Protein electron-transfer, Proteins, Scanning tunneling microscopy, Silver halides, Single molecule, Single molecules

Quantum dots (QDs) are semiconductor nanoparticles with unique optical and electronic properties, whose interest as potential nano-theranostic platforms for imaging and sensing is increasing. The design and use of QDs requires the understanding of cell-nanoparticle interactions at a microscopic and nanoscale level. Model systems such as supported lipid bilayers (SLBs) are useful, less complex platforms mimicking physico-chemical properties of cell membranes. In this work, we investigated the effect of topographical homogeneity of SLBs bearing different surface charge in the adsorption of hydrophilic QDs. Using quartz-crystal microbalance, a label-free surface sensitive technique, we show significant differences in the interactions of QDs onto homogeneous and inhomogeneous SLBs formed following different strategies. Within short time scales, QDs adsorb onto topographically homogeneous, defect-free SLBs is driven by electrostatic interactions, leading to no layer disruption. After prolonged QD exposure, the nanomechanical stability of the SLB decreases suggesting nanoparticle insertion. In the case of inhomogeneous, defect containing layers, QDs target preferentially membrane defects, driven by a subtle interplay of electrostatic and entropic effects, inducing local vesicle rupture and QD insertion at membrane edges. © 2021

JTD Keywords: adsorption, atomic force microscopy, bilayer formation, gold nanoparticles, hydrophilic quantum dots, lipid membrane defects, model, nanomechanics, quartz crystal microbalance with dissipation, size, supported lipid bilayers, surfaces, Atomic force microscopy, Atomic-force-microscopy, Cytology, Defect-free, Electronic properties, Electrostatics, Hydrophilic quantum dot, Hydrophilic quantum dots, Hydrophilicity, Hydrophilics, Lipid bilayers, Lipid membrane defect, Lipid membrane defects, Lipid membranes, Lipids, Nanocrystals, Nanomechanics, Optical and electronic properties, Quartz, Quartz crystal microbalance with dissipation, Quartz crystal microbalances, Quartz-crystal microbalance, Semiconductor nanoparticles, Semiconductor quantum dots, Supported lipid bilayers

The use of silver nanoparticles (Ag NPs) in the biomedical field deserves a mindful analysis of the possible inflammatory response which could limit their use in the clinic. Despite the anti-cancer properties of Ag NPs having been widely demonstrated, there are still few studies concerning their involvement in the activation of specific inflammatory pathways. The inflammatory outcome depends on the synthetic route used in the NPs production, in which toxic reagents are employed. In this work, we compared two types of Ag NPs, obtained by two different chemical routes: conventional synthesis using sodium citrate and a green protocol based on leaf extracts as a source of reduction and capping agents. A careful physicochemical characterization was carried out showing spherical and stable Ag NPs with an average size between 20 nm and 35 nm for conventional and green Ag NPs respectively. Then, we evaluated their ability to induce the activation of inflammation in Human Leukemic Monocytes (THP-1) differentiated into M0 macrophages using 1 µM and 2 µM NPs concentrations (corresponded to 0.1 µg/mL and 0.2 µg/mL respectively) and two-time points (24 h and 48 h). Our results showed a clear difference in Nuclear Factor ?B (NF-?b) activation, Interleukins 6–8 (IL-6, IL-8) secretion, Tumor Necrosis Factor-? (TNF-?) and Cyclooxygenase-2 (COX-2) expression exerted by the two kinds of Ag NPs. Green Ag NPs were definitely tolerated by macrophages compared to conventional Ag NPs which induced the activation of all the factors mentioned above. Subsequently, the exposure of breast cancer cell line (MCF-7) to the green Ag NPs showed that they exhibited antitumor activity like the conventional ones, but surprisingly, using the MCF-10A line (not tumoral breast cells) the green Ag NPs did not cause a significant decrease in cell viability. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

JTD Keywords: activation, biosynthesis, gold nanoparticles, green route, inflammation response, mechanism, metal, nanotechnology, physico-chemical properties, raman-spectroscopy, resonance, silver nanoparticles, surface, Biomedical fields, Cell culture, Cell death, Chemical activation, Chemical routes, Conventional synthesis, Diseases, Green route, Inflammation response, Inflammatory response, Macrophages, Metal nanoparticles, Nf-kappa-b, Pathology, Physico-chemical properties, Physicochemical property, Property, Silver nanoparticles, Sodium compounds, Synthetic routes, Toxic reagents

The SARS-CoV-2 outbreaks highlighted the need for effective, reliable, fast, easy-to-do and cheap diagnostics procedures. We pragmatically experienced that an early positive-case detection, inevitably coupled with a mass vaccination campaign, is a milestone to control the COVID-19 pandemic. Gold nanoparticles (AuNPs) can indeed play a crucial role in this context, as their physicochemical, optics and electronics properties are being extensively used in photothermal therapy (PTT), radiation therapy (RT), drug delivery and diagnostic. AuNPs can be synthesized by several approaches to obtain different sizes and shapes that can be easily functionalized with many kinds of molecules such as antibodies, proteins, probes, and lipids. In addition, AuNPs showed high biocompatibility making them useful tool in medicine field. We thus reviewed here the most relevant evidence on AuNPs as effective way to detect the presence of SARS-CoV-2 antigens. We trust future diagnostic efforts must take this 'old-fashioned' nanotechnology tool into consideration for the development and commercialization of reliable and feasible detection kits.

JTD Keywords: Aggregation, Antibodies, Assay, Covid-19, Diagnosis, Enhanced raman-scattering, Gold nanoparticles, Immunoassay, Pandemic disease, Physicochemical properties, Rapid detection, Sars-cov-2, Sensors, Surface-plasmon resonance, Therapy

Contact-based antimicrobials, as antibiotic-free technologies that use non-specific interactions with bacterial cells to exert antimicrobial activity, are a prospective solution in fighting the global issue of bacterial resistance. A very simplified approach to their design considers the direct bonding of cationic guanidine-containing amino acids to the surface of nano-gold carriers. The structure enables antimicrobial activity due to a high density of cationic surface charges. This opens a set of novel questions that are important for their effective engineering, particularly regarding (i) chemistry and events that take place at the interface between NPs and cells, (ii) the direct influence of a charge (and its change) on interactions with bacterial and mammalian cells, and (iii) the stability of structures (and their antimicrobial activity) in the presence of enzymes, which are addressed in this paper. Because of the ability of amino acid-functionalized nano-gold to retain structural and functional activity, even after exposure to a range of physicochemical stimuli, they provide an excellent nanotechnological platform for designing highly effective contact-based antimicrobials and their applications.

JTD Keywords: agents, antibiotic-free technology, arginine, charged amino acids, contact-based antimicrobials, discovery, enzyme-resistant antimicrobials, functionalized gold, peptides, polymers, resistant, Antibiotic-free technology, Charged amino acids, Contact-based antimicrobials, Enzyme-resistant antimicrobials, Functionalized gold, Nanoparticles

© 2020 Wiley-VCH GmbH Cytosolic protein delivery remains elusive. The inability of most proteins to cross the cellular membrane is a huge hurdle. Here we explore the unique photothermal properties of gold nanorods (AuNRs) to trigger cytosolic delivery of proteins. Both partners, protein and AuNRs, are modified with a protease-resistant cell-penetrating peptide with nuclear targeting properties to induce internalization. Once internalized, spatiotemporal control of protein release is achieved by near-infrared laser irradiation in the safe second biological window. Importantly, catalytic amounts of AuNRs are sufficient to trigger cytosolic protein delivery. To the best of our knowledge, this is the first time that AuNRs with their maximum of absorption in the second biological window are used to deliver proteins into the intracellular space. This strategy represents a powerful tool for the cytosolic delivery of virtually any class of protein.

JTD Keywords: cell-penetrating peptide, cytosolic delivery, gold nanorod, near-infrared irradiation, Cell-penetrating peptide, Cytosolic delivery, Gold nanorod, Near-infrared irradiation

© 2020 Elsevier Ltd The use of nanoparticles in consumer products is currently on the rise, so it is important to have reliable methods to predict any associated toxicity effects. Traditional in vitro assays fail to mimic true physiological responses of living organisms against nanoparticles whereas murine in vivo models are costly and ethically controversial. For these reasons, this study aimed to evaluate the efficacy of Galleria mellonella as an alternative, non-rodent in vivo model for examining nanoparticle toxicity. Silver, selenium, and functionalized gold nanoparticles were synthesized, and their toxicity was assessed in G. mellonella larvae. The degree of acute toxicity effects caused by each type of NP was efficiently detected by an array of indicators within the larvae: LD50 calculation, hemocyte proliferation, NP distribution, behavioral changes, and histological alterations. G. mellonella larvae are proposed as a nanotoxicological model that can be used as a bridge between in vitro and in vivo murine assays in order to obtain better predictions of NP toxicity.

JTD Keywords: cellular uptake, cytotoxicity, galleria mellonella, gold nanoparticles, hemocytes, nanoparticles, nanotoxicity, non-rodent in vivo model, non-rodent in vivo model, oxidative stress, selenium-compounds, silica nanoparticles, silver nanoparticles, toxicity, toxicity screening, vitro, Galleria mellonella, Hemocytes, In-vivo model, Nanoparticles, Nanotoxicity, Non-rodent in vivo model, Toxicity screening

© 2021 American Chemical Society. Photosynthetic reactions in plants, algae, and cyanobacteria are driven by photosystem I and photosystem II complexes, which specifically reduce or oxidize partner redox biomolecules. Photosynthetic complexes can also bind synthetic organic molecules, which inhibit their photoactivity and can be used both to study the electron transport chain and as herbicides and algicides. Thus, their development, characterization, and sensing bears fundamental and applied interest. Substantial efforts have been devoted to developing photosensors based on photosystem II to detect compounds that bind to the plastoquinone sites of this complex. In comparison, photosystem I based sensors have received less attention and could be used to identify novel substances displaying phytotoxic effects, including those obtained from natural product extracts. We have developed a robust procedure to functionalize gold electrodes with photo- and redox-active photosystem I complexes based on transparent gold and a thiolate self-assembled monolayer, and we have obtained reproducible electrochemical photoresponses. Chronoamperometric recordings have allowed us to measure photocurrents in the presence of the viologen derivative paraquat at concentrations below 100 nM under lock-in operation and a sensor dynamic range spanning six orders of magnitude up to 100 mM. We have modeled their time course to identify the main electrochemical processes and limiting steps in the electron transport chain. Our results allow us to isolate the contributions from photosystem I and the redox mediator, and evaluate photocurrent features (spectral and power dependence, fast transient kinetics) that could be used as a sensing signal to detect other inhibitors and modulators of photosystem I activity.

JTD Keywords: biosensor, herbicide, kinetic model, paraquat, photo-chrono-amperometry, photosystem i, self-assembled monolayer, transparent gold microelectrode, Biosensor, Herbicide, Kinetic model, Paraquat, Photo-chrono-amperometry, Photosystem i, Self-assembled monolayer, Transparent gold microelectrode

© 2020 The Authors Although the concept of selective delivery has been postulated over 100 years ago, no targeted nanomedicine has been clinically approved so far. Nanoparticles modified with targeting ligands to promote the selective delivery of therapeutics towards a specific cell population have been extensively reported. However, the rational design of selective particles is still challenging. One of the main reasons for this is the lack of quantitative theoretical and experimental understanding of the interactions involved in cell targeting. In this review, we discuss new theoretical models and experimental methods that provide a quantitative view of targeting. We show the new advancements in multivalency theory enabling the rational design of super-selective nanoparticles. Furthermore, we present the innovative approaches to obtain key targeting parameters at the single-cell and single molecule level and their role in the design of targeting nanoparticles. We believe that the combination of new theoretical multivalent design and experimental methods to quantify receptors and ligands aids in the rational design and clinical translation of targeted nanomedicines.

JTD Keywords: binding-kinetics, biological identity, biomolecular corona, blood-brain-barrier, drug-delivery, gold nanoparticles, multivalency, nanotechnology, protein corona, quantitative characterization, rational design, super-selectivity, superresolution microscopy, tumor heterogeneity, Ligand-receptor interactions, Multivalency, Nanotechnology, Quantitative characterization, Rational design, Super-selectivity

Engineered immunoglobulin-G molecules (IgGs) are of wide interest for the development of detection elements in protein-based biosensors with clinical applications. The strategy usually employed for the de novo design of such engineered IgGs consists on merging fragments of the three-dimensional structure of a native IgG, which is immobilized on the biosensor surface, and of an antibody with an exquisite target specificity and affinity. In this work conventional and accelerated classical molecular dynamics (cMD and aMD, respectively) simulations have been used to propose two IgG-like antibodies for COVID-19 detection. More specifically, the crystal structure of the IgG1 B12 antibody, which inactivates the human immunodeficiency virus-1, has been merged with the structure of the antibody CR3022 Fab tightly bounded to SARS-CoV-2 receptor-binding domain (RBD) and the structure of the 5309 antibody Fab fragment complexed with SARS-CoV-2 RBD. The two constructed antibodies, named IgG1-CR3022 and IgG1-S309, respectively, have been immobilized on a stable gold surface through a linker. Analyses of the influence of both the merging strategy and the substrate on the stability of the two constructs indicate that the IgG1-S309 antibody better preserves the neutralizing structure than the IgG1-CR3022 one. Overall, results indicate that the IgG1-S309 is appropriated for the generation of antibody based sensors for COVID-19 diagnosis. (C) 2021 The Author(s). Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology.

JTD Keywords: cr3022, igg1, molecular engineering, s309, Antibodies, Antibody engineering, Biosensors, Chemical detection, Clinical application, Cov, Cr3022, Crystal structure, Design, Diseases, Gold nanoparticles, Igg1, Igg1 antibody, Immobilization, Immunoglobulin g, Immunosensor, In-silico, Merging, Molecular dynamics, Molecular engineering, Orientation, Protein-based biosensors, Receptor-binding domains, S309, Sars, Sensor, Spike protein, Target, Vaccine, Viruses

Background: Emerging concepts for designing innovative drugs (i.e., novel generations of antimicrobials) frequently include nanostructures, new materials, and nanoparticles (NPs). Along with numerous advantages, NPs bring limitations, partly because they can limit the analytical techniques used for their biological and in vivo validation. From that standpoint, designing innovative drug delivery systems requires advancements in the methods used for their testing and investigations. Considering the well-known ability of resazurin-based methods for rapid detection of bacterial metabolisms with very high sensitivity, in this work we report a novel optimization for tracking bacterial growth kinetics in the presence of NPs with specific characteristics, such as specific optical properties. Results: Arginine-functionalized gold composite (HAp/Au/arginine) NPs, used as the NP model for validation of the method, possess plasmonic properties and are characterized by intensive absorption in the UV/vis region with a surface plasmon resonance maximum at 540 nm. Due to the specific optical properties, the NP absorption intensively interferes with the light absorption measured during the evaluation of bacterial growth (optical density; OD600). The results confirm substantial nonspecific interference by NPs in the signal detected during a regular turbidity study used for tracking bacterial growth. Instead, during application of a resazurin-based method (Presto Blue), when a combination of absorption and fluorescence detection is applied, a substantial increase in the signal-to-noise ratio is obtained that leads to the improvement of the accuracy of the measurements as verified in three bacterial strains tested with different growth rates (E. coli, P. aeruginosa, and S. aureus). Conclusions: Here, we described a novel procedure that enables the kinetics of bacterial growth in the presence of NPs to be followed with high time resolution, high sensitivity, and without sampling during the kinetic study. We showed the applicability of the Presto Blue method for the case of HAp/Au/arginine NPs, which can be extended to various types of metallic NPs with similar characteristics. The method is a very easy, economical, and reliable option for testing NPs designed as novel antimicrobials.

JTD Keywords: Antimicrobial nanoparticles, Arginine-functionalized gold, Bacterial growth kinetics, Plasmonic nanoparticles, Presto Blue

The inclusion compound (IC) of cyclodextrin (CD) containing the antitumor drug Methotrexate (MTX) as a guest molecule was obtained to increase the solubility of MTX and decrease its inherent toxic effects in nonspecific cells. The IC was conjugated with gold nanoparticles (AuNPs), obtained by a chemical method, creating a ternary intelligent delivery system for MTX molecules, based on the plasmonic properties of the AuNPs. Irradiation of the ternary system, with a laser wavelength tunable with the corresponding surface plasmon of AuNPs, causes local energy dissipation, producing the controlled release of the guest from CD cavities. Finally, cell viability was evaluated using MTS assays for β-CD/MTX and AuNPs + β-CD/MTX samples, with and without irradiation, against HeLa tumor cells. The irradiated sample of the ternary system AuNPs + β-CD/MTX produced a diminution in cell viability attributed to the photothermal release of MTX.

JTD Keywords: Cyclodextrin, Delivery system, Gold nanoparticles, Inclusion compound, Irradiation, Laser, Methotrexate, Photothermal release

JTD Keywords: DNA nanotechnology, Gold nanoparticles, Lithography, Metasurfaces, Nanopatterning

We report the first DNA-based origami technique that can print addressable patterns on surfaces with sub-10 nm resolution. Specifically, we have used a two-dimensional DNA origami as a template (DNA origami stamp) to transfer DNA with pre-programmed patterns (DNA ink) on gold surfaces. The DNA ink is composed of thiol-modified staple strands incorporated at specific positions of the DNA origami stamp to create patterns upon thiol-gold bond formation on the surface (DNA ink). The DNA pattern formed is composed of unique oligonucleotide sequences, each of which is individually addressable. As a proof-of-concept, we created a linear pattern of oligonucleotide-modified gold nanoparticles complementary to the DNA ink pattern. We have developed an in silico model to identify key elements in the formation of our DNA origami-driven lithography and nanoparticle patterning as well as simulate more complex nanoparticle patterns on surfaces.

JTD Keywords: DNA nanotechnology, Lithography, Nanopatterning, Gold nanoparticles, Metasurfaces

Evidence of guest migration in α-cyclodextrin-octylamine (α-CD-OA) inclusion compound (IC) generated via plasmonic heating of gold nanoparticles (AuNPs) has been studied. In this report, we demonstrate local effects generated by laser-mediated irradiation of a sample of AuNPs covered with inclusion compounds on surface-derivatized glass under liquid conditions by atomic force microscopy (AFM). Functionalized AuNPs on the glass and covered by the ICs were monitored by recording images by AFM during 5 h of irradiation, and images showed that after irradiation, a drastic decrease in the height of the AuNPs occurred. The absorption spectrum of the irradiated sample showed a hypsochromic shift from 542 to 536 nm, evidence suggesting that much of the population of nanoparticles lost all of the parts of the overlay of ICs due to the plasmonic heat generated by the irradiation. Mass spectrometry matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) performed on a sample containing a collection of drops obtained from the surface of the functionalized glass provided evidence that the irradiation lead to disintegration of the ICs and therefore exit of the octylamine molecule (the guest) from the cyclodextrin cavity (the matrix).

JTD Keywords: Cyclodextrin inclusion compound, Gold nanoparticles, Guest migration, Plasmonic heating

We show that optical visualization of ultrathin mica flakes on metallic substrates is viable using semitransparent gold as substrates. This enables to easily localize mica flakes and rapidly estimate their thickness directly on gold substrates by conventional optical reflection microscopy. We experimentally demonstrate it by comparing optical images with atomic force microscopy images of mica flakes on semitransparent gold. Present results open the possibility for simple and rapid characterization of thin mica flakes as well as other thin sheets directly on metallic substrates.

JTD Keywords: Atomic force, Conductive AFM, Gold substrates, Metallic substrate, Optical image, Optical reflection, Optical visualization, Ultrathin layers, Atomic force microscopy, Geometrical optics, Gold, Mica, Optical microscopy, Substrates

Calcium phosphate compounds can potentially influence cellular fate through ionic substitutions. However, to be able to turn such solution-mediated processes into successful directors of cellular response, a perfect understanding of the material-induced chemical reactions in situ is required. We therefore report on the application of home-made electrochemical microelectrodes, tested as pH and chloride sensors, for precise spatial and temporal characterization of different aqueous environments around calcium phosphate-based biomaterials prepared from α-tricalcium phosphate using clinically relevant liquid to powder ratios. The small size of the electrodes allowed for online measurements in traditionally inaccessible in vitro environments, such as the immediate material-liquid interface and the interior of curing bone cement. The kinetic data obtained has been compared to theoretical sorption models, confirming that the proposed setup can provide key information for improved understanding of the biochemical environment imposed by chemically reactive biomaterials.

JTD Keywords: Calcium phosphate, Hydroxyapatite, Ion sorption, Iridium oxide, Sensors, Animals, Biocompatible Materials, Bone Cements, Calcium Phosphates, Cells, Cultured, Chlorides, Electrochemical Techniques, Gold, Hydrogen-Ion Concentration, Hydroxyapatites, Iridium, Materials Testing, Microelectrodes, Powders, Silver, Silver Compounds, Water

We demonstrate the possibility to manipulate 25 nm radius gold nanoparticles in the three spatial dimensions with an atomic force microscope with the use of electroenhanced capillary forces. We show that an enhanced water-bridge can be electrostatically induced between a conducting probe and a metallic nanoparticle by the application of a voltage pulse, which is able to exert a pulling capillary force on the nanoparticle strong enough to detach it from the substrate. The nanoparticle can then be moved, attached to the probe, and placed back to the desired location on the substrate simply by contacting it.

JTD Keywords: Atomic force microscopy, Capillarity, Gold, Nanoparticles, Nanotechnology

In this work, we report the detection of proteins by means of simultaneous fluorescence and impedance measurements in a cyclo olefin polymer (COP) chip containing an ordered pillar array column, used for reversed-phase liquid chromatography, with integrated microband gold electrodes at the end of the channel.

JTD Keywords: Cyclo olefin polymer, Gold microelectrodes, Impedance, Pillar array, Protein detection

In this study, we compare for the first time the nanomechanical properties of lipid bilayer islands on flat and porous surfaces. 1,2-Dimyzistoyl-sn-glycero-3-phosphatidylcholine (DMPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) bilayers were deposited on flat (silicon and mica) and porous silicon (pSi) substrate surfaces and examined using atomic force spectroscopy and force volume imaging. Force spectroscopy measurements revealed the effects of the underlying substrate and of the lipid phase on the nanomechanical properties of bilayers islands. For mica and silicon, significant differences in breakthrough force between the center and the edges of bilayer islands were observed for both phospolipids. These differences were more pronounced for DMPC than for DPPC, presumably due to melting effects at the edges of DMPC bilayers. In contrast, bilayer islands deposited on pSi yielded similar breakthrough forces in the central region and along the perimeter of the islands, and those values in turn were similar to those measured along the perimeter of bilayer islands deposited on the flat substrates. The study also demonstrates that pSi is suitable solid support for the formation of pore-spanning phospholipid bilayers with potential applications in transmembrane protein studies, drug delivery, and biosensing.

JTD Keywords: Black lipid-membranes, Gold surfaces, Supported bilayers, Channel activity, Micro-BLMS, Silicon, Proteins, Vesicles, AFM, Temperature measurement

This work describes our studies on the molecular design of interfacial architectures suitable for DNA sensing which could resist non-specific binding of nanomaterials commonly used as labels for amplifying biorecognition events. We observed that the non-specific binding of bio-nanomaterials to surface-confined oligonucleotide strands is highly dependent on the characteristics of the interfacial architecture. Thiolated double stranded oligonucleotide arrays assembled on Au surfaces evidence significant fouling in the presence of nanoparticles (NPs) at the nanomolar level. The non-specific interaction between the oligonucleotide strands and the nanomaterials can be sensitively minimized by introducing streptavidin (SAv) as an underlayer conjugated to the DNA arrays. The role of the SAv layer was attributed to the significant hydrophilic repulsion between the SAv-modified surface and the nanomaterials in close proximity to the interface, thus conferring outstanding anti-fouling characteristics to the interfacial architecture. These results provide a simple and straightforward strategy to overcome the limitations introduced by the non-specific binding of labels to achieve reliable detection of DNA-based biorecognition events.

JTD Keywords: DNA/ analysis, Gold, Nanostructures/ chemistry, Oligonucleotide Array Sequence Analysis/ instrumentation, Oligonucleotides/ chemistry, Streptavidin/ chemistry, Sulfhydryl Compounds

The mechanical properties of alkanethiol monolayers on Au(111) in KOH solution have been studied by force spectroscopy. The analysis of the vertical force versus penetration curves showed that monolayer penetration is a stepped process that combines elastic regions with sudden penetration events. The structural meaning of these events can be explained both by the creation of gauche defects on the hydrocarbon chains and by a cooperative molecular tilting model proposed by Barrena et al. [J. Chem. Phys. 113, 2413 (2000)]. The validity of these models for alkanethiol monolayers of different compactness and chain length has been discussed. The Young's modulus (E) of the monolayers has been calculated by using a recently developed model which considers the thickness of the monolayer as a parameter, thus allowing a decoupling of the mechanical properties of the thiol layer from those of the Au(111) substrate. As a result, the calculated E values are in the range of 50-150 Pa, which are remarkably lower than those previously reported in the literature.

JTD Keywords: Adsorbed layers, AFM, Gold, Monolayers, Organic compounds, Self-assemblyYoung's modulus