Enhanced oligonucleotide-nanoparticle conjugate stability using thioctic acid modified oligonucleotides

Metallic nanoparticles of gold functionalized with oligonucleotides conventionally use a terminal thiol modification and have been used in a wide range of applications. Although readily available, the oligonucleotide–nanoparticle conjugates prepared in this way suffer from a lack of stability when exposed to a variety of small molecules or elevated temperatures. If silver is used in place of gold then this lack of stability is even more pronounced. In this study we report the synthesis of highly stabilized oligonucleotide–nanoparticle conjugates using a simple oligonucleotide modification. A modified solid support was used to generate 3′-thioctic acid modified oligonucleotides by treatment with an N-hydroxysuccimidyl ester of thioctic acid. Unusually, both gold and silver nanoparticles have been investigated in this study and show that these disulphide-modified oligonucleotide probes offer significant improvements in nanoparticle stability when treated with dithiothreitol (DTT) compared with monothiol analogues. This is a significant advance in oligonucleotide–nanoparticle conjugate stability and for the first time allows silver nanoparticles to be prepared that are more stable than standard gold-thiol functionalized nanoparticles. This opens up the possibility of using silver nanoparticles functionalized with oligonucleotides as an alternative to gold.     

Sucrose density gradient centrifugation separation of gold and silver nanoparticles synthesized using Magnolia kobus plant leaf extracts

The synthesis and post-synthesis separation of nanoparticles that are polydispersed in size and shape is important due to their variety of applications. In the present study, it is demonstrated that the Magnolia kobus plant extract produces a diverse mixture of extracellular gold and silver nanocrystals with a majority of polydispersed spheres; however, there are a significant number of homogeneously sized triangles, pentagons, and hexagons. The gold and silver nanoparticles synthesized using the M. kobus plant extract can be separated using density gradient centrifugation in the size range of 52 ～ 117 nm and 38 ～ 61 nm, respectively. The average particle sizes increase with increases in the sucrose concentration of each layer. Relatively larger but long, thin plates of gold nanoparticles appear in the higher density sediments, whereas a larger proportion of smaller spheres featured in the lower density gradients. Similarly, silver nanospheres of different sizes are separated at different density gradients with smaller proportions of plates.     

Green synthesis of gold nanoparticles with Zingiber officinale extract: Characterization and blood compatibility

Biosynthesis of gold nanoparticles with small size and biostability is very important and used in various biomedical applications. There are lot of reports for the synthesis of gold nanoparticles by the addition of reducing agent and stabilizing agent. In the present study we have synthesized gold nanoparticles, with a particle size ranging from 5 to 15 nm, using Zingiber officinale extract which acts both as reducing and stabilizing agent. Z. officinale extract is reported to be a more potent anti-platelet agent than aspirin. Therefore, green synthesis of gold nanoparticles with Z. officinale extract, as an alternative to chemical synthesis, is beneficial from its biological and medical applications point of view, because of its good blood biocompatibility and physiological stability. The formation and size distribution of gold nanoparticles were confirmed by dynamic light scattering (DLS), UV–vis spectrophotometer and transmission electron microscopy (TEM). Gold nanoparticles synthesized using citrate and Z. officinale extract demonstrated very low protein adsorption. Both nanoparticles were non platelet activating and non complement activating on contact with whole human blood. They also did not aggregate other blood cells, however, nanoparticles synthesised with Z. officinale extract was highly stable at physiological condition compared to citrate capped nanoparticles, which aggregated. Thus the usage of nanoparticles, synthesized with Z. officinale extract, as vectors for the applications in drug delivery, gene delivery or as biosensors, where a direct contact with blood occurs is justified.     

Potential of plant as a biological factory to synthesize gold and silver nanoparticles and their applications

Green synthesis of metallic nanoparticles has become a promising field of research in recent years. Syntheses of gold and silver nanoparticles by various chemical and physical methods as well as the biosynthetic approach mediated by numerous microorganisms have been actively researched. A more scalable and economic route to produce these metallic nanoparticles would be through the plant-mediated synthetic approach. Owing to the biodiversity of plant biomasses, the mechanism by which bioconstituents of plants have contributed to the synthetic process is yet to be fully understood. Nevertheless, the feasibility of controlling the shape and size of nanoparticles by varying the reaction conditions has been demonstrated in many studies. This paper provides an overview of the plant-mediated syntheses of gold and silver nanoparticles, possible compounds and mechanisms that might be responsible for the bioreduction process as well as the potential applications of biosynthesized nanoparticles in different fields. The challenges and limitations of this plant-mediated biosynthetic approach are also discussed.     

Starch vermicelli template-assisted synthesis of size/shape-controlled nanoparticles

Size- and shape-controlled syntheses of silver and gold nanoparticles have been successfully developed using partially hydrolyzed starch vermicelli templates as green nanoreactors for the growth of nanoparticles. Mung bean vermicelli is of interest due to the higher amylose content and its transparency, allowing the formation of coloured particles on the vermicelli to be observed. The as-prepared silver and gold nanoparticles were characterized by UV–Visible spectroscopy, transmission electron microscopy (TEM), and X-ray diffraction (XRD). The carbonization of as-prepared vermicelli at 200 °C, 300 °C, and 500 °C was carried out to investigate nanoparticles embedded in the starch vermicelli templates. TEM of carbonized samples revealed the interesting patterns of gold nanorods and silver nanowire-liked assemblies along with carbon nanotubes. The carbonization of silver nanoparticles at 500 °C resulted to the loss of starch vermicelli capping nanoparticles and this led to the higher diffusion rate of nanoparticles to generate silver nanodendrites on TEM images. XRD data of carbonized yellow and purple silver nanoparticles revealed the presence of silver nanoparticles and a mixture of silver and silver chloride nanoparticles, respectively. This approach offers a great potential to design new fine structures of vermicelli and utilize its structure as a template for the large-scale synthesis of size- and shape-controlled silver and gold nanoparticles for chemical and biological applications.     

Silver and Gold Nanoparticles Exposure to In Vitro Cultured Retina – Studies on Nanoparticle Internalization,Apoptosis, Oxidative Stress,Glial- and Microglial Activity

The complex network of neuronal cells in the retina makes it a potential target of neuronal toxicity – a risk factor for visual loss. With growing use of nanoparticles (NPs) in commercial and medical applications, including ophthalmology, there is a need for reliable models for early prediction of NP toxicity in the eye and retina. Metal NPs, such as gold and silver, gain much of attention in the ophthalmology community due to their potential to cross the barriers of the eye. Here, NP uptake and signs of toxicity were investigated after exposure to 20 and 80 nm Ag- and AuNPs, using an in vitro tissue culture model of the mouse retina. The model offers long-term preservation of retinal cell types, numbers and morphology and is a controlled system for delivery of NPs, using serum-free defined culture medium. AgNO₃-treatment was used as control for toxicity caused by silver ions. These end-points were studied; gross morphological organization, glial activity, microglial activity, level of apoptosis and oxidative stress, which are all well described as signs of insult to neural tissue. TEM analysis demonstrated cellular- and nuclear uptake of all NP types in all neuronal layers of the retina. Htx-eosin staining showed morphological disruption of the normal complex layered retinal structure, vacuole formation and pyknotic cells after exposure to all Ag- and AuNPs. Significantly higher numbers of apoptotic cells as well as an increased number of oxidative stressed cells demonstrated NP-related neuronal toxicity. NPs also caused increased glial staining and microglial cell activation, typical hallmarks of neural tissue insult. This study demonstrates that low concentrations of 20 and 80 nm sized Ag- and AuNPs have adverse effects on the retina, using an organotypic retina culture model. Our results motivate careful assessment of candidate NP, metallic or-non-metallic, to be used in neural systems for therapeutic approaches.     

Optimization of immunolabeled plasmonic nanoparticles for cell surface receptor analysis

Noble metal nanoparticles hold great potential as optical contrast agents due to a unique feature, known as the plasmon resonance, which produces enhanced scattering and absorption at specific frequencies. The plasmon resonance also provides a spectral tunability that is not often found in organic fluorophores or other labeling methods. The ability to functionalize these nanoparticles with antibodies has led to their development as contrast agents for molecular optical imaging. In this review article, we present methods for optimizing the spectral agility of these labels. We discuss synthesis of gold nanorods, a plasmonic nanoparticle in which the plasmonic resonance can be tuned during synthesis to provide imaging within the spectral window commonly utilized in biomedical applications. We describe recent advances in our group to functionalize gold and silver nanoparticles using distinct antibodies, including EGFR, HER-2 and IGF-1, selected for their relevance to tumor imaging. Finally, we present characterization of these nanoparticle labels to verify their spectral properties and molecular specificity.     

Optimization of Nanoparticle Size for Plasmonic Enhancement of Fluorescence

This paper reports on the enhancement of fluorescence that can result from the proximity of fluorophores to metallic nanoparticles (NPs). This plasmonic enhancement, which is a result of the localized surface plasmon resonance at the metal surface, can be exploited to improve the signal obtained from optical biochips and thereby lower the limits of detection. There are two distinct enhancement effects: an increase in the excitation of the fluorophore and an increase in its quantum efficiency. This study focuses on the first of these effects where the maximum enhancement occurs when the NP plasmon resonance wavelength coincides with the fluorophore absorption band. In this case, the excitation enhancement is proportional to the square of the amplitude of the electric field. The scale of the enhancement depends on many parameters, such as NP size and shape, metal type, and NP–fluorophore separation. A model system consisting of spherical gold/silver alloy NPs, surrounded by a silica spacer shell, to which is attached a fluorescent ruthenium dye, was chosen and the dependence of the fluorescence enhancement on NP diameter was investigated. Theoretical calculations, based on Mie theory, were carried out to predict the maximum possible enhancement factor for spherical NPs with a fixed composition and a range of diameters. Spherical NPs of the same composition were fabricated by chemical preparation techniques. The NPs were coated with a thin silica shell to overcome quenching effects and the dye was attached to the shell.     

Nanosilver—The burgeoning therapeutic molecule and its green synthesis

Nanotechnology is a field that is burgeoning day by day, making an impact in all spheres of human life. Biological methods of synthesis have paved way for the “greener synthesis” of nanoparticles and these have proven to be better methods due to slower kinetics, they offer better manipulation and control over crystal growth and their stabilization. This has motivated an upsurge in research on the synthesis routes that allow better control of shape and size for various nanotechnological applications. Nanosilver has developed as a potent antibacterial, antifungal, anti-viral and anti-inflammatory agent. The recent advancement in the field includes the enzymatic method of synthesis suggesting enzymes to be responsible for the nanoparticle formation. The biomedical applications of silver nanoparticle can be effective by the use of biologically synthesized nanoparticles which minimize the factors such as toxicity and cost and are found to be exceptionally stable. The targeting of cancer cells using silver nanoparticles has proven to be effective, but neither the exact mechanism of action nor the modes of activation of the downstream signaling molecules have been revealed yet. The review illustrates a probable signaling pathway and mechanism by which silver nanoparticles target the cancer cells. The current review also examines the historical background of nanoparticles, role of silver nanoparticles in various biomedical applications and also focusing on better methods of the synthesis of nanoparticles.     

Electrogenerated chemiluminescence of luminol in neutral and alkaline aqueous solutions on a silver nanoparticle self-assembled gold electrode.

The electrochemiluminescence (ECL) behaviour of luminol on a silver nanoparticle self-assembled gold electrode in neutral and alkaline solutions was investigated using conventional cyclic voltammetry (CV). The silver nanoparticle self-assembled gold electrode exhibited excellent ECL properties for the luminol ECL system. In neutral solutions, four ECL peaks (ECL-1-ECL-4) were observed at 0.73, 1.15, -0.46 and -1.35 V (vs. SCE), respectively. The intensities of these peaks were enhanced significantly compared with those on a bulk gold electrode and a gold nanoparticle self-assembled gold electrode. It was found that ECL-1 and ECL-2 on a silver nanoparticle-modified electrode were about 1000 and 1770 times stronger than those on a bare Au electrode and were about 17 and 15 times stronger than those on a gold nanoparticle-modified electrode, respectively. In alkaline solutions, four ECL peaks were also observed that were much stronger than those in neutral solutions, and ECL-1 and ECL-2 were enhanced by about three orders and one order of magnitude compared with those on a bare Au electrode and on a gold nanoparticle self-assembled electrode, respectively. Moreover, the silver nanoparticle-modified electrode exhibited good stability and reproducibility for luminol ECL. These peaks were found to depend on a number of factors, including silver nanoparticles on the surface of the modified electrode, potential scan direction, scan rate, scan range, the presence of O2 or N2, pH values, the concentrations of NaBr and luminol, and buffer solutions. The emitter of the ECL was confirmed as 3-aminophthalate by analysing the CL spectra. The surface state of the silver nanoparticle self-assembled electrode was characterized by scanning electron microscopy (SEM) and the interface property of the electrode was studied by electrochemical impedance spectroscopy (EIS). A mechanism for the formation of these ECL peaks is proposed. The results demonstrate that luminol has excellent ECL properties, such as strong ECL intensity and good reproducibility on a silver nanoparticle-modified gold electrode, in both neutral and alkaline solutions, which is of great potential in analytical applications.     

A review on biological synthesis of ZnO nanostructures and its application in photocatalysis mediated dye degradation: An overview

Zinc oxide (ZnO) has several industrial applications due to its versatile properties, which lead to its continuously increasing demand in different industrial sectors. Additionally, ZnO nanostructures possess unique photocatalytic activity, and because of this, they are being applied to degrade organic dyes through photocatalysis for wastewater treatment. Nevertheless, chemical synthesis methods to develop ZnO nanostructures have raised concerns related to environmental issues, furthermore, these methods are found to be costly and tedious. As a result, the synthesis of ZnO nanostructures using green methods is gaining popularity due to its low cost and eco-friendly mode, while avoiding the use of toxic chemicals. Green synthesis of ZnO nanostructures using different biological approaches involving plants, algae, and different microorganism-derived bioactive compounds has been well reported for diverse applications. Among different applications, ZnO nanostructures that enable photocatalysis to degrade dye have been found to be imperative for wastewater treatments. Therefore, the current review explores recent studies on green synthesis approaches to prepare ZnO nanostructures via adopting different biological methods that rely on plants, algae, and bacterial microorganisms. The properties of ZnO nanostructures, along with their green synthesis routes and feasible mechanisms, have also been discussed in this review. This review focuses on the use and efficiency of green route synthesized ZnO nanostructures as nanophotocatalysts for the degradation of organic dyes in wastewater treatment. Additionally, existing challenges in green synthesis methods and the efficiency of ZnO nanostructures to degrade organic dyes following photocatalysis has been discussed.     

Tansy fruit mediated greener synthesis of silver and gold nanoparticles

In this paper we have reported the green synthesis of silver (AgNPs) and gold (AuNPs) nanoparticles by reduction of silver nitrate and chloroauric acid solutions, respectively, using fruit extract of Tanacetum vulgare; commonly found plant in Finland. The process for the synthesis of AgNPs and AuNPs is rapid, novel and ecofriendly. Formation of the AgNPs and AuNPs were confirmed by surface plasmon spectra using UV–Vis spectrophotometer and absorbance peaks at 452 and 546 nm. Different tansy fruit extract concentration (TFE), silver and gold ion concentration, temperature and contact times were experimented in the synthesis of AgNPs and AuNPs. The properties of prepared nanoparticles were characterized by TEM, XRD, EDX and FTIR. Finally zeta potential values at various pH were analyzed along with corresponding SPR spectra.     

Synthesis of polysaccharide-stabilized gold and silver nanoparticles: a green method

A simple, green method was developed for the synthesis of gold and silver nanoparticles by using polysaccharides as reducing/stabilizing agents. The obtained positively charged chitosan-stabilized gold nanoparticles and negatively charged heparin-stabilized silver nanoparticles were characterized with UV-vis spectroscopy and transmission electron microscopy. The results illustrated the formation of gold and silver nanoparticles inside the nanoscopic polysaccharide templates. Moreover, the morphology and size distribution of prepared gold and silver nanoparticles varied with the concentration of both the polysaccharides and the precursor metal salts.     

Biological synthesis of gold nanowires using extract of Rhodopseudomonas capsulata

An environmentally friendly method using a cell-free extract (CFE) of Rhodopseudomonas capsulata is proposed to synthesize gold nanowires with a network structure. This procedure offers control over the shapes of gold nanoparticles with the change of HAuCl4 concentration. The CFE solutions were added with different concentrations of HAuCl4, resulting in the bioreduction of gold ions and biosynthesis of morphologies of gold nanostructures. It is probable that proteins acted as the major biomolecules involved in the bioreduction and synthesis of gold nanoparticles. At a lower concentration of gold ions, exclusively spherical gold nanoparticles with sizes ranging from 10 to 20 nm were produced, whereas gold nanowires with a network structure formed at the higher concentration of gold ions in the aqueous solution. This method is expected to be applicable to the synthesis of other metallic nanowires such as silver and platinum, and even other anisotropic metal nanostructures are expected using the biosynthetic methods.     

Biological synthesis of nanoparticles in biofilms

The biological synthesis of nanoparticles (NPs) by bacteria and biofilms via extracellular redox reactions has received attention because of the minimization of harmful chemicals, low cost, and ease of culturing and downstream processing. Bioreduction mechanisms vary across bacteria and growth conditions, which leads to various sizes and shapes of biosynthesized NPs. NP synthesis in biofilms offers additional advantages, such as higher biomass concentrations and larger surface areas, which can lead to more efficient and scalable biosynthesis. Although biofilms have been used to produce NPs, the mechanistic details of NP formation are not well understood. In this review, we identify three critical areas of research and development needed to advance our understanding of NP production by biofilms: 1) synthesis, 2) mechanism and 3) stabilization. Advancement in these areas could result in the biosynthesis of NPs that are suitable for practical applications, especially in drug delivery and biocatalysis. Specifically, the current status of methods and mechanisms of nanoparticle synthesis and surface stabilization using planktonic bacteria and biofilms is discussed. We conclude that the use of biofilms to synthesize and stabilize NPs is underappreciated and could provide a new direction in biofilm-based NP production.     

Gold@Silver@Gold Core Double-Shell Nanoparticles: Synthesis and Aggregation-Enhanced Two-Photon Photoluminescence Evaluation

A facile, straightforward, and low-cost method is proposed to synthesize gold@silver@gold core double-shell nanoparticles. The technique is a seed-mediated growth protocol that contains four steps of (1) gold seed synthesis, (2) gold seed growth, (3) silver layer coating through silver salt reduction, and (4) gold layer deposition via gold precursor reduction. The prepared nanoparticles had a narrow size distribution and the average particle size of 28 ± 1 nm. Cysteine was introduced to the nanoparticles solution as a coupling agent to assemble nanoparticles. Aggregation-induced two-photon photoluminescence enhancement of three types assembled nanoparticles, i.e., gold@silver@gold, gold@silver, and gold nanoparticles, was studied. It was observed that the assembled core double-shell nanoparticles presented huge enhancement in two-photon photoluminescence signal in comparison with two other nanoparticles. Moreover, the gold@silver@gold nanoparticle is a stable and biocompatible plasmonic nanosystem. This paper provides a novel candidate for two-photon photoluminescence excitation sensing and imaging for biomedical applications.

     

Investigation of immunogold-silver staining by electron microscopy

Deposition of metallic silver on colloidal gold immunoreagents has been shown to be a very sensitive immunostaining technique capable of detecting low levels of immunoreactivity in tissue sections. Using electron microscopy we have shown that immunolabelling is highest with small sizes of gold which can penetrate sections better and achieve higher densities of particles in the section than larger particles. Chemical permeabilisation of the embedding medium aids the penetration of colloidal gold. The silver enhancement step in immunogold-silver staining was shown to be progressive, allowing optimisation of staining and the selection of the final size of silver deposits required. Some poorly understood features of the technique are rationalised and the additional knowledge gained will aid the wider application of this method.     

Silver-enhanced colloidal gold as a cell surface marker for photoelectron microscopy

Colloidal gold labeling in conjunction with silver enhancement was investigated as a labeling technique for photoelectron microscopy (PEM). PEM uses UV-stimulated electron emission to image uncoated cell surfaces, and markers for cell surfaces need to be sufficiently photoemissive to be clearly visible against this background. Label contrast provided by 6 nm or 20 nm colloidal gold markers alone was compared to that provided by 6 nm markers after silver enhancement, using both direct and indirect labeling methods for fibronectin on human fibroblast cell surfaces. In all cases, details of the fibrillar fibronectin labeling distribution which were barely discernible before silver enhancement became highly visible against the cellular surface features. Two factors evidently contribute to the pronounced increase in label contrast with silver enhancement: (1) Increased particle size, which was documented by transmission electron microscopy, and (2) increased photoemission resulting from a silver coating on the enhanced gold markers, compared with the protein coating on the unenhanced gold markers. These data demonstrate that silver enhancement of colloidal gold labeling patterns in PEM images is a highly effective method for localization of specific sites on cell surfaces.     

Gas-Aggregated Copper Nanoparticles with Long-term Plasmon Resonance Stability

Metal nanoparticles (NPs) possessing localized surface plasmon resonance (LSPR) are of high interest for applications in optics, electronics, catalysis, and sensing. The practically important issue is the stability of the LSPR, which often limits the use of some metals due to their chemical reactivity leading to degradation of the NP functionality. In this work, copper NPs of two distinct sizes are produced by magnetron sputtering gas aggregation. This method ensures formation of the particles with high purity and monocrystallinity, enhancing the chemical inertness and providing a superior time stability of the plasmonic properties. Additionally, a simple UV-ozone treatment, which leads to the formation of an oxide shell around the copper NPs, is found to be an efficient method to prevent following gradual oxidation and assure the LSPR stability in ambient atmospheric conditions for periods over 100 days even for small (10–12 nm in diameter) NPs. The obtained results allow for significant improvement of the competitiveness of copper NPs with gold or silver nanostructures, which are traditionally used in plasmonics.

     

Exploitation of marine bacteria for production of gold nanoparticles

Background

Gold nanoparticles (AuNPs) have found wide range of applications in electronics, biomedical engineering, and chemistry owing to their exceptional opto-electrical properties. Biological synthesis of gold nanoparticles by using plant extracts and microbes have received profound interest in recent times owing to their potential to produce nanoparticles with varied shape, size and morphology. Marine microorganisms are unique to tolerate high salt concentration and can evade toxicity of different metal ions. However, these marine microbes are not sufficiently explored for their capability of metal nanoparticle synthesis. Although, marine water is one of the richest sources of gold in the nature, however, there is no significant publication regarding utilization of marine micro-organisms to produce gold nanoparticles. Therefore, there might be a possibility of exploring marine bacteria as nanofactories for AuNP biosynthesis.

Results

In the present study, marine bacteria are exploited towards their capability of gold nanoparticles (AuNPs) production. Stable, monodisperse AuNP formation with around 10?nm dimension occur upon exposure of HAuCl₄ solution to whole cells of a novel strain of Marinobacter pelagius, as characterized by polyphasic taxonomy. Nanoparticles synthesized are characterized by Transmission electron microscopy, Dynamic light scattering and UV-visible spectroscopy.

Conclusion

The potential of marine organisms in biosynthesis of AuNPs are still relatively unexplored. Although, there are few reports of gold nanoparticles production using marine sponges and sea weeds however, there is no report on the production of gold nanoparticles using marine bacteria. The present work highlighted the possibility of using the marine bacterial strain of Marinobacter pelagius to achieve a fast rate of nanoparticles synthesis which may be of high interest for future process development of AuNPs. This is the first report of AuNP synthesis by marine bacteria.