Interactions Between Plasmonic Nanostructures and Proteins

In the development of a nanodevice for biomedical applications, the study of the interactions with the biomolecules is essential. Proteins, in particular, are known to be easily adsorbed on the surface of the nanoparticles and the resulting complex is the one that will be effectively internalized by the target cells. Owing to the versatility of the preparation methods available and the unique optical properties, gold nanomaterials represent an excellent choice to study this interaction. This chapter will initially describe the synthesis of gold nanorods and nanoshells that are able to absorb light in the near-infrared (NIR) region. Then, the methods available for the functionalization of their surface will be discussed. The surface plasmon absorption will be used as an optical tool to monitor the process of preparation and surface modification. In the last section of the chapter, fluorescence and microscopy techniques will be used to follow the formation and characterize the protein-nanoparticle complex. The modifications of the emission spectra of two model proteins, bovine serum albumin (BSA) and myoglobin (Mb), will be analyzed in detail. The data will demonstrate that structural rearrangements following the adsorption on the surface of the nanoparticles are responsible for the changes in the fluorescence of the tryptophan residues of the protein. The data will be discussed in terms of static and dynamic quenching, proving the formation of a protein-nanoparticle complex. Atomic force microscopy (AFM) measurements will allow the direct visualization of this complex

UV Treatment of the Stabilizing Shell for Improving the Photostability of Silver Nanoparticles

Silver nanoparticles or nanoclusters are quite sensitive to light exposure. In particular, irradiation in the localized surface plasmon resonance (LSPR) region brings about a drastic modification of their optical properties due to growth and reshaping of the nanoparticles. In order to obtain luminescent colloids, small silver colloidal nanoparticles were prepared in chloroform using vinylpyrrolidone oligomers as capping agent and their luminescence properties were used to control their stability upon prolonged exposure to visible light. The polymeric shell around the metal clusters was hardened through photo-cross-linking by UV light. This process did not alter the morphology and the optical properties of the nanoparticles but greatly improved the particle photostability as confirmed also by confocal laser scanning microscopy measurements. The data clearly show that UV curing of the stabilizing layer could be a simple postsynthetic procedure to obtain materials with stable properties

Controlled assembly of metal colloids on dye-doped silica particles to tune the photophysical properties of organic molecules

The use of plasmonic nanomaterials is a challenging strategy to control radiation and radiation-induced processes at a nanometric scale. The localized surface plasmons of metal nanoparticles have been shown to affect the efficiency of a variety of radiative and non-radiative processes occurring in organic molecules. In this contribution, we present an overview of the results obtained through an original approach based on the hierarchical assembly of plasmonic gold colloids on silica templates, covalently doped with organic dyes. The detailed morphological characterization demonstrates the disposition of gold colloids on silica achieved through the tight control of the synthetic conditions. The studies carried out while gradually increasing the concentration of gold nanoparticles allow the detailed investigation of the effects of the progressive addition of plasmonic particles on the photophysical behaviour of organic molecules. In particular, the fluorescence behaviour of three dyes with different spectral properties, namely fluorescein, rhodamine B and 9-aminoacridine, are investigated in the presence of increasing concentrations of gold nanoparticles. In order to fix the distance between the dye and the gold nanoparticles, the dyes are anchored to silica nanoparticles, and the metal colloids are chemically adsorbed on the silica surface. The steady state and time-resolved data are analysed to evaluate the impact of plasmonic nanoparticles on the radiative and non-radiative processes of the dyes; the data provide evidence that the modulation of the fluorescence intensity (enhancement or quenching) can be achieved by changing the concentration of gold colloids. The plasmonic nanostructures can be employed to favour one deactivation process over the others. For example, we demonstrate that the photoinduced formation of reactive oxygen species (ROS) can be enhanced upon the plasmonic engineering of a photosensitizing agent (Protoporphyrin IX, PpIX). The Vis-excitation of silica-PpIX samples in the presence of gold nanoparticles results in a faster and more efficient photoinduced formation of ROS species either in solution or in a hydrogel. The ROS efficiency data and the fluorescence behaviour of PpIX in the presence of gold colloids suggest that the enhancement of the excitation field occurs through a plasmonic effect. For the application of the assembled hybrid materials, further advantages come from the development of photosensitizer-containing hydrogel films that are able to efficiently produce ROS upon visible excitation. Our preliminary results are herein reported and discussed

photoluminescence of a single quantum emitter in a strongly inhomogeneous chemical environment

A comprehensive photoluminescence study of defect centers in single SiO2 nanoparticles provides new insight into the complex photo-physics of single quantum emitters embedded into a random chemical environment

Driving the Interactions between Organic Nanoparticles and Phospolipidic Membranes by an Easy Treatment of the Surface Stabilizer

Polymer-stabilized perylene nanoparticles were prepared through a solvent exchange method. The formation of the nanostructures in aqueous solution was confirmed by the appearance of a red-shifted emission attributable to the formation of excimer-like aggregates. The behavior of organic nanostructures in the presence of lipid vesicles was investigated through steady-state and time-resolved fluorescence measurements. When no further surface treatment is applied to the nanoparticles, changes in the decay times and emission spectra demonstrate that inside the lipid bilayers the nanoparticles redissolve into the monomeric form with a rate and efficiency determined by the working temperature (above and below the transition temperature <i>T</i>_m of the phospholipid). On the other hand, when the stabilized shell is UV-cured to induce photo-cross-linking of the polymeric chains, the nanoparticle stability increases and their redissolution in the membrane is prevented. Confocal fluorescence images support the data obtained in bulk. The results indicate that the prepared nanostructures could be successfully used either as nanometric carriers for the delivery of poor water-soluble lipophilic compounds or as imaging tools depending on the rigidity/cross-linking degree of their polymeric stabilizer shell

Selective internalization of ZnAl-HTlc nanoparticles in normal and tumor cells. A study of their potential use in cellular delivery

A colloidal dispersion of zinc aluminum hydrotalcite nanoparticles (ZnAl-HTlc) has been used for in vitro experimental procedures in order to provide reliable data on their potential application in cellular delivery. Two different cell lines (HeLa tumor cells and MDCK normal cells) with a similar epithelial derivation have been used. Sedimentation studies performed in the presence of different constituents of the cell culture medium revealed the importance of serum components to stabilize the colloidal dispersions of nanosized ZnAl-HTlc. Cell viability assay showed for nanosized ZnAl-HTlc a higher cell growth inhibition on tumor cells compared to normal cells whereas LDH test showed the absence of toxicity for both cell lines. Cellular uptake experiments indicated a preferential internalization of ZnAl-HTlc nanoparticles in HeLa tumor cells. Adsorption study and steady state fluorescence measurements on the phenol red/HTlc hybrid were carried out in order to verify the possibility of using phenol red as fluorescent dye for ZnAl-HTlc nanoparticles. The observed spectral behavior indicated a strong interaction between the dye and the inorganic matrix and the preferential adsorption of the dye on the nanoparticle surface has been confirmed by the XRPD data. Fluorescence confocal imaging showed a different localization pattern of nanosized HTlc in the two cell lines and a higher fluorescence signals in tumor cells supporting the occurrence of more efficient internalization processes in the pathogen cell line as observed in the cellular uptake experiments. © 2011 Elsevier B.V

The Influence of Modified Silica Nanomaterials on Adult Stem Cell Culture

The preparation of tailored nanomaterials able to support cell growth and viability is mandatory for tissue engineering applications. In the present work, silica nanoparticles were prepared by a sol-gel procedure and were then functionalized by condensation of amino groups and by adsorption of silver nanoparticles. Transmission electron microscopy (TEM) imaging was used to establish the morphology and the average dimensions of about 130 nm, which were not affected by the functionalization. The three silica samples were deposited (1 mg/mL) on cover glasses, which were used as a substrate to culture adult human bone marrow-mesenchymal stem cells (hBM-MSCs) and human adipose-derived stem cells (hASCs). The good cell viability over the different silica surfaces was evaluated by monitoring the mitochondrial dehydrogenase activity. The analysis of the morphological parameters (aspect ratio, cell length, and nuclear shape Index) yielded information about the interactions of stem cells with the surface of three different nanoparticles. The data are discussed in terms of chemical properties of the surface of silica nanoparticles

Spectroscopic and Microscopic Studies of Aggregation and Fibrillation of Lysozyme in Water/Ethanol Solutions

The thermal aggregation of lysozyme has been analyzed in water/ethanol solutions at low pH to induce the specific protein aggregation pathway which leads to fibrillar structures in a few hours. In this solvating medium, the protein undergoes a conformational rearrangement promoting the formation of fibrils that are structurally similar to amyloid ones. As the process evolves with different steps, a multitechnique approach has been used by means of analytical probes that can be selectively sensitive in the detection of the different stages of protein association. Fourier transform infrared spectroscopy, intrinsic fluorescence, stationary fluorescence anisotropy, transmission electron microscopy (TEM), and atomic force microscopy (AFM) measurements have been carried out at different times to access and characterize the whole aggregation pathway. The data recorded with different experimental setups revealed different sensitivity to different stages of protein assembling. The whole set of data together with the direct visualization of different aggregate structures by use of TEM and AFM imaging enable to discuss a possible mechanism of fibrillation