Anti-aquaporin-4 immunoglobulin G colorimetric detection by silver nanoparticles.

Neuromyelitis optica spectrum disorder (NMOSD) is an inflammatory and autoimmune disease whose biomarker is the anti-AQP4-IgG autoantibody that binds to aquaporin-4 (AQP4) protein. We introduced a nanosensor with a sensitivity of 84.6%, higher than the CBA's 76.5%. Using silver nanoparticles (AgNPs), we detected not only seropositive but also some false-negative patients previously classified with CBA. It consisted of AgNPs coated with one of a panel of 5 AQP4 epitopes. The ability in detecting the anti-AQP4-IgG in NMOSD patients depended on the epitope and synergy could be obtained by combining different epitopes. We demonstrated that NMOSD patients could easily be distinguished from healthy subjects and patients with multiple sclerosis. Using the most sensitive AQP461-70 peptide, we established a calibration curve to estimate the concentration of anti-AQP4-IgG in seropositive NMOSD patients. The ability to enhance the accuracy of the diagnosis may improve the prognosis of 10-27% of anti-AQP4-IgG seronegative patients worldwide.

A New Dermal Substitute Containing Polyvinyl Alcohol with Silver Nanoparticles and Collagen with Hyaluronic Acid: In Vitro and In Vivo Approaches.

The experimental use of poly (alcohol-vinyl) (PVA) as a skin curative is increasing widely. However, the use of this hydrogel is challenging due to its favorable properties for microbiota growth. The association with silver nanoparticles (AgNPs) as an antimicrobial agent turns the match for PVA as a dressing, as it focuses on creating a physical barrier to avoid wound dehydration. When associated with extracellular components, such as the collagen matrix, the device obtained can create the desired biological conditions to act as a skin substitute. This study aimed to analyze the anti-microbiological activity and the in vitro and in vivo responses of a bilaminar device of PVA containing AgNPs associated with a membrane of collagen-hyaluronic acid (col-HA). Additionally, mesenchymal stem cells were cultured in the device to evaluate in vitro responses and in vivo immunomodulatory and healing behavior. The device morphology revealed a porous pattern that favored water retention and in vitro cell adhesion. Controlled wounds in the dorsal back of rat skins revealed a striking skin remodeling with new epidermis fulfilling all previously injured areas after 14 and 28 days. No infections or significant inflammations were observed, despite increased angiogenesis, and no fibrosis-markers were identified as compared to controls. Although few antibacterial activities were obtained, the addition of AgNPs prevented fungal growth. All results demonstrated that the combination of the components used here as a dermal device, chosen according to previous miscellany studies of low/mid-cost biomaterials, can promote skin protection avoiding infections and dehydration, minimize the typical wound inflammatory responses, and favor the cellular healing responses, features that give rise to further clinical trials of the device here developed.

Peptide-Conjugated Silver Nanoparticle for Autoantibody Recognition.

In this work, we considered the autoantibodies proposed as putative biomarkers of demyelination taking into account their reactivity towards myelin oligodendrocyte glycoprotein (MOG) and myelin basic protein (MBP). These myelin proteins are among the most commonly researched targets in the immunopathology of demyelinating diseases. In this context, the development of assays for autoantibody detection can contribute as a predictive value for the early diagnosis of demyelinating diseases. Hence, we aimed to address the application of silver nanoparticles (AgNPs) as a sensing device of autoantibodies. AgNPs were synthesized via a chemical reduction method and characterized using atomic force microscopy (AFM), X-ray diffractometry, dynamic light scattering, and UV-visible spectrophotometry. The process of peptide conjugation on the nanoparticles was also analyzed. The autoantibody recognition by the peptide-conjugated AgNPs was evaluated with UV-visible spectrophotometry, atomic force spectroscopy (AFS), and color changing. AgNPs exhibited spherical morphology, low polydispersity, face-centered cubic crystal structure, and an average size of 29.3±3.0 nm. The hydrodynamic diameter variation and AFM showed that the MBP peptide induced greater agglomeration, compared to MOG peptide. The AFS measurements indicated the efficient binding of peptides to the AgNPs maintaining their activity, revealed by typical adhesion force and shapes of curves. The absorption spectrum features were more affected by the interaction with the specific autoantibodies, which also caused a visible color change in suspension providing a qualitative response. We described a preliminary study of MOG- and MBP-conjugated AgNPs which showed to be applicable in the autoantibody recognition. These have promising implication in the searching for biological markers for diagnostic purposes in the demyelination context, in which the nanoscale sensing exploitation is recent.

Cantilever nanobiosensor using tyrosinase to detect atrazine in liquid medium.

The aim of this study was to develop a cantilever nanobiosensor for atrazine detection in liquid medium by immobilising the biological recognition element (tyrosinase vegetal extract) on its surface with self-assembled monolayers using gold, 16-mercaptohexadecanoic acid, 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride/n-hydroxysuccinimide. Cantilever nanobiosensors presented a surface compression tension increase when atrazine concentrations were increased, with a limit of detection and limit of quantification of 7.754 ppb (parts per billion) and 22.792 ppb, respectively. From the voltage results obtained, the evaluation of atrazine contamination in river and drinking water were very close to those of the reference sample and ultrapure water, demonstrating the ability of the cantilever nanobiosensor to distinguish different water samples and different concentrations of atrazine. Cantilever nanosensor surface functionalization was characterised by combining polarisation modulation infrared reflection-absorption spectroscopy and atomic force microscopy and indicating film thickness in nanometric scale (80.2 ± 0.4 nm). Thus, the cantilever nanobiosensor developed for this study using low cost tyrosinase vegetal extract was adequate for atrazine detection, a potential tool in the environmental field.

Adsorption according to the Langmuir-Freundlich model is the detection mechanism of the antigen p53 for early diagnosis of cancer.

Biosensors for early detection of cancer biomarkers normally depend on specific interactions between such biomarkers and immobilized biomolecules in the sensing units. Though these interactions are expected to yield specific, irreversible adsorption, the underlying mechanism appears not to have been studied in detail. In this paper, we show that adsorption explained with the Langmuir-Freundlich model is responsible for detection of the antigen p53 associated with various types of cancers. Irreversible adsorption was proven between anti-p53 antibodies immobilized on the biosensors and the antigen p53, with the adequacy of the Langmuir-Freundlich model being confirmed with three independent experimental methods, viz. polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS), nanogravimetry using a quartz crystal microbalance and electrochemical impedance spectroscopy. The method based on this irreversible adsorption was sufficiently sensitive (limit of detection of 1.4 pg mL(-1)) for early diagnosis of Hodgkin lymphoma, pancreatic and colon carcinomas, and bladder, ovarian and lung cancers, and could distinguish between MCF7 cells containing the antigen p53 from Saos-2 cells that do not contain it.

Modeling the coverage of an AFM tip by enzymes and its application in nanobiosensors.

A stochastic simulation of adsorption processes was developed to simulate the coverage of an atomic force microscope (AFM) tip with enzymes represented as rigid polyhedrons. From geometric considerations of the enzyme structure and AFM tip, we could estimate the average number of active sites available to interact with substrate molecules in the bulk. The procedure was exploited to determine the interaction force between acetyl-CoA carboxylase enzyme (ACC enzyme) and its substrate diclofop, for which steered molecular dynamics (SMD) was used. The theoretical force of (1.6±0.5) nN per enzyme led to a total force in remarkable agreement with the experimentally measured force with AFM, thus demonstrating the usefulness of the procedure proposed here to assist in the interpretation of nanobiosensors experiments.

Nanobiosensors based on chemically modified AFM probes: a useful tool for metsulfuron-methyl detection.

The use of agrochemicals has increased considerably in recent years, and consequently, there has been increased exposure of ecosystems and human populations to these highly toxic compounds. The study and development of methodologies to detect these substances with greater sensitivity has become extremely relevant. This article describes, for the first time, the use of atomic force spectroscopy (AFS) in the detection of enzyme-inhibiting herbicides. A nanobiosensor based on an atomic force microscopy (AFM) tip functionalised with the acetolactate synthase (ALS) enzyme was developed and characterised. The herbicide metsulfuron-methyl, an ALS inhibitor, was successfully detected through the acquisition of force curves using this biosensor. The adhesion force values were considerably higher when the biosensor was used. An increase of ~250% was achieved relative to the adhesion force using an unfunctionalised AFM tip. This considerable increase was the result of a specific interaction between the enzyme and the herbicide, which was primarily responsible for the efficiency of the nanobiosensor. These results indicate that this methodology is promising for the detection of herbicides, pesticides, and other environmental contaminants.

Theoretical models for surface forces and adhesion and their measurement using atomic force microscopy.

The increasing importance of studies on soft matter and their impact on new technologies, including those associated with nanotechnology, has brought intermolecular and surface forces to the forefront of physics and materials science, for these are the prevailing forces in micro and nanosystems. With experimental methods such as the atomic force spectroscopy (AFS), it is now possible to measure these forces accurately, in addition to providing information on local material properties such as elasticity, hardness and adhesion. This review provides the theoretical and experimental background of afs, adhesion forces, intermolecular interactions and surface forces in air, vacuum and in solution.

Atomic force microscopy as a tool applied to nano/biosensors.

This review article discusses and documents the basic concepts and principles of nano/biosensors. More specifically, we comment on the use of Chemical Force Microscopy (CFM) to study various aspects of architectural and chemical design details of specific molecules and polymers and its influence on the control of chemical interactions between the Atomic Force Microscopy (AFM) tip and the sample. This technique is based on the fabrication of nanomechanical cantilever sensors (NCS) and microcantilever-based biosensors (MC-B), which can provide, depending on the application, rapid, sensitive, simple and low-cost in situ detection. Besides, it can provide high repeatability and reproducibility. Here, we review the applications of CFM through some application examples which should function as methodological questions to understand and transform this tool into a reliable source of data. This section is followed by a description of the theoretical principle and usage of the functionalized NCS and MC-B technique in several fields, such as agriculture, biotechnology and immunoassay. Finally, we hope this review will help the reader to appreciate how important the tools CFM, NCS and MC-B are for characterization and understanding of systems on the atomic scale.

Low-cost gas sensors produced by the graphite line-patterning technique applied to monitoring banana ripeness.

A low-cost sensor array system for banana ripeness monitoring is presented. The sensors are constructed by employing a graphite line-patterning technique (LPT) to print interdigitated graphite electrodes on tracing paper and then coating the printed area with a thin film of polyaniline (PANI) by in-situ polymerization as the gas-sensitive layer. The PANI layers were used for the detection of volatile organic compounds (VOCs), including ethylene, emitted during ripening. The influence of the various acid dopants, hydrochloric acid (HCl), methanesulfonic acid (MSA), p-toluenesulfonic acid (TSA) and camphorsulfonic acid (CSA), on the electrical properties of the thin film of PANI adsorbed on the electrodes was also studied. The extent of doping of the films was investigated by UV-Vis absorption spectroscopy and tests showed that the type of dopant plays an important role in the performance of these low-cost sensors. The array of three sensors, without the PANI-HCl sensor, was able to produce a distinct pattern of signals, taken as a signature (fingerprint) that can be used to characterize bananas ripeness.

Designing an enzyme-based nanobiosensor using molecular modeling techniques.

Nanobiosensors can be built via functionalization of atomic force microscopy (AFM) tips with biomolecules capable of interacting with the analyte on a substrate, and the detection being performed by measuring the force between the immobilized biomolecule and the analyte. The optimization of such sensors may require multiple experiments to determine suitable experimental conditions for the immobilization and detection. In this study we employ molecular modeling techniques to assist in the design of nanobiosensors to detect herbicides. As a proof of principle, the properties of acetyl co-enzyme A carboxylase (ACC) were obtained with molecular dynamics simulations, from which the dimeric form in an aqueous solution was found to be more suitable for immobilization owing to a smaller structural fluctuation than the monomeric form. Upon solving the nonlinear Poisson-Boltzmann equation using a finite-difference procedure, we found that the active sites of ACC exhibited a positive surface potential while the remainder of the ACC surface was negatively charged. Therefore, optimized biosensors should be prepared with electrostatic adsorption of ACC onto an AFM tip functionalized with positively charged groups, leaving the active sites exposed to the analyte. The preferential orientation for the herbicides diclofop and atrazine with the ACC active site was determined by molecular docking calculations which displayed an inhibition coefficient of 0.168 µM for diclofop, and 44.11 µM for atrazine. This binding selectivity for the herbicide family of diclofop was confirmed by semiempirical PM6 quantum chemical calculations which revealed that ACC interacts more strongly with the herbicide diclofop than with atrazine, showing binding energies of -119.04 and +8.40 kcal mol(-1), respectively. The initial measurements of the proposed nanobiosensor validated the theoretical calculations and displayed high selectivity for the family of the diclofop herbicides.

Synthesis of nanoparticles and nanofibers of polyaniline by potentiodynamic electrochemical polymerization.

Potentiodynamic electrochemical synthesis was used to controllably synthesize nanofibers (mean diameter 48 nm) and/or nanoparticles (mean diameter 88 nm) of polyaniline (PANI) on gold electrodes. The films were characterized by cyclic voltammetry (CV), field emission gun scanning electron microscopy (FEG-SEM) and atomic force microscopy (AFM). The type and dimensions of the nanostructures depend on deposition conditions such as monomer concentration and scan rate. This study shows that the nucleation and growth steps play a key role on the film development and its nano-morphology.

Nanoscale conformational ordering in polyanilines investigated by SAXS and AFM.

Understanding the adsorption mechanisms in nanostructured polymer films has become crucial for their use in technological applications, since film properties vary considerably with the experimental conditions utilized for film fabrication. In this paper, we employ small-angle X-ray scattering (SAXS) to investigate solutions of polyanilines and correlate the chain conformations with morphological features of the nanostructured films obtained with atomic force microscopy (AFM). It is shown that aggregates formed already in solution affect the film morphology; in particular, at early stages of adsorption film morphology appears entirely governed by the chain conformation in solution and adsorption of aggregates. We also use SAXS data for modeling poly(o-ethoxyaniline) (POEA) particle shape through an ab initio procedure based on simulated annealing using the dummy atom model (DAM), which is then compared to the morphological features of POEA films fabricated with distinct pHs and doping acids. Interestingly, when the derivative POEA is doped with p-toluene sulfonic acid (TSA), the resulting films exhibit a fibrillar morphology-seen with atomic force microscopy and transmission electron microscopy-that is consistent with the cylindrical shape inferred from the SAXS data. This is in contrast with the globular morphology observed for POEA films doped with other acids.