Exocytosis of peptide functionalized gold nanoparticles in endothelial cells

We present the exocytosis profile of two types of peptide-coated nanoparticles, which have similar charge and size but different functionality. While one kind of particles appears to progressively exocytose, the other one has a more complex profile, suggesting that some of the particles are re-uptaken by the cells. Both types of particles retain their colloidal stability after exocytosis

Total synthesis of isotopically enriched Si-29 silica NPs as potential spikes for isotope dilution quantification of natural silica NPs

A new method was developed for the preparation of highly monodisperse isotopically enriched Si-29 silica nanoparticles (29Si-silica NPs) with the purpose of using them as spikes for isotope dilution mass spectrometry (IDMS) quantification of silica NPs with natural isotopic distribution. Si-29 tetraethyl orthosilicate (29Si-TEOS), the silica precursor was prepared in two steps starting from elementary silicon-29 pellets. In the first step Si-29 silicon tetrachloride (29SiCl4) was prepared by heating elementary silicon-29 in chlorine gas stream. By using a multistep cooling system and the dilution of the volatile and moisture-sensitive 29SiCl4 in carbon tetrachloride as inert medium we managed to reduce product loss caused by evaporation. 29Si-TEOS was obtained by treating 29SiCl4 with absolute ethanol. Structural characterisation of 29Si-TEOS was performed by using 1H and 13C nuclear magnetic resonance (NMR) spectroscopy and Fourier-transform infrared (FTIR) spectroscopy. For the NP preparation, a basic amino acid catalysis route was used and the resulting NPs were analysed using transmission electron microscopy (TEM), small angle X-ray scattering (SAXS), dynamic light scattering (DLS) and zeta potential measurements. Finally, the feasibility of using enriched NPs for on-line field-flow fractionation coupled with multi-angle light scattering and inductively coupled plasma mass spectrometry (FFF/MALS/ICP-MS) has been demonstrated

A comparison of techniques for size measurement of nanoparticles in cell culture medium

Plain and aminated silica nanoparticles dispersed in purified water, in 50 mM Tris-HCl buffer and in cell culture medium were measured using dynamic light scattering (DLS), centrifugal liquid sedimentation (CLS), small-angle X-ray scattering (SAXS), and particle tracking analysis (PTA). The test samples were measured by all methods immediately after dispersion and after incubation at room temperature for 24 h. The effect of the biological dispersion medium on the modal value of the particle size distribution was compared for each method taking into account the estimated uncertainty. For the methods based on light scattering, DLS and PTA, the size distributions obtained were significantly altered due to the formation of a protein corona and induced agglomeration effects. With SAXS and CLS, the measured size of the primary particles was mostly unchanged. While SAXS offers excellent precision and traceability to the SI unit system if the model fitting approach is used for data analysis, CLS provides detailed size distributions from which additional information on the agglomeration state can be deduced

Synthesis of Porous Hollow Organosilica Particles with Tunable Shell Thickness

Porous hollow silica particles possess promising applications in many fields, ranging from drug delivery to catalysis. From the synthesis perspective, the most challenging parameters are the monodispersity of the size distribution and the thickness and porosity of the shell of the particles. This paper demonstrates a facile two-pot approach to prepare monodisperse porous-hollow silica particles with uniform spherical shape and well-tuned shell thickness. In this method, a series of porous-hollow inorganic and organic-inorganic core-shell silica particles were synthesized via hydrolysis and condensation of 1,2-bis(triethoxysilyl) ethane (BTEE) and tetraethyl orthosilicate (TEOS) in the presence of hexadecyltrimethylammonium bromide (CTAB) as a structure-directing agent on solid silica spheres as core templates. Finally, the core templates were removed via hydrothermal treatment under alkaline conditions. Transmission electron microscopy (TEM) was used to characterize the particles′ morphology and size distribution, while the changes in the chemical composition during synthesis were followed by Fourier-transform infrared spectroscopy. Single-particle inductively coupled plasma mass spectrometry (spICP-MS) was applied to assess the monodispersity of the hollow particles prepared with different reaction parameters. We found that the presence of BTEE is key to obtaining a well-defined shell structure, and the increase in the concentration of the precursor and the surfactant increases the thickness of the shell. TEM and spICP-MS measurements revealed that fused particles are also formed under suboptimal reaction parameters, causing the broadening of the size distribution, which can be preceded by using appropriate concentrations of BTEE, CTAB, and ammonia

Versailles project on advanced materials and standards (VAMAS) interlaboratory study on measuring the number concentration of colloidal gold nanoparticles

We describe the outcome of a large international interlaboratory study of the measurement of particle number concentration of colloidal nanoparticles, project 10 of the technical working area 34, "Nanoparticle Populations" of the Versailles Project on Advanced Materials and Standards (VAMAS). A total of 50 laboratories delivered results for the number concentration of 30 nm gold colloidal nanoparticles measured using particle tracking analysis (PTA), single particle inductively coupled plasma mass spectrometry (spICP-MS), ultraviolet-visible (UV-Vis) light spectroscopy, centrifugal liquid sedimentation (CLS) and small angle X-ray scattering (SAXS). The study provides quantitative data to evaluate the repeatability of these methods and their reproducibility in the measurement of number concentration of model nanoparticle systems following a common measurement protocol. We find that the population-averaging methods of SAXS, CLS and UV-Vis have high measurement repeatability and reproducibility, with between-labs variability of 2.6%, 11% and 1.4% respectively. However, results may be significantly biased for reasons including inaccurate material properties whose values are used to compute the number concentration. Particle-counting method results are less reproducibile than population-averaging methods, with measured between-labs variability of 68% and 46% for PTA and spICP-MS respectively. This study provides the stakeholder community with important comparative data to underpin measurement reproducibility and method validation for number concentration of nanoparticles

Versailles project on advanced materials and standards (VAMAS) interlaboratory study on measuring the number concentration of colloidal gold nanoparticles

We describe the outcome of a large international interlaboratory study of the measurement of particle number concentration of colloidal nanoparticles, project 10 of the technical working area 34, "Nanoparticle Populations" of the Versailles Project on Advanced Materials and Standards (VAMAS). A total of 50 laboratories delivered results for the number concentration of 30 nm gold colloidal nanoparticles measured using particle tracking analysis (PTA), single particle inductively coupled plasma mass spectrometry (spICP-MS), ultraviolet-visible (UV-Vis) light spectroscopy, centrifugal liquid sedimentation (CLS) and small angle X-ray scattering (SAXS). The study provides quantitative data to evaluate the repeatability of these methods and their reproducibility in the measurement of number concentration of model nanoparticle systems following a common measurement protocol. We find that the population-averaging methods of SAXS, CLS and UV-Vis have high measurement repeatability and reproducibility, with between-labs variability of 2.6%, 11% and 1.4% respectively. However, results may be significantly biased for reasons including inaccurate material properties whose values are used to compute the number concentration. Particle-counting method results are less reproducibile than population-averaging methods, with measured between-labs variability of 68% and 46% for PTA and spICP-MS respectively. This study provides the stakeholder community with important comparative data to underpin measurement reproducibility and method validation for number concentration of nanoparticles

Workshop on Regulatory Preparedness for Innovation in Nanotechnology

This report summarises the presentations and discussions at the first NanoReg2 Workshop on Regulatory Preparedness for Innovation in Nanotechnology held in Ispra, Italy 5 to 6 October 2017 and attended by approximately 60 regulators, industry representatives and other stakeholders. NanoReg2 is a European Union (EU) Horizon 2020 project. At the workshop, Regulatory Preparedness was defined as the regulators' timely awareness of innovations and the regulator's actions to check whether present legislation covers all safety aspects of each innovation, including initiating revision of the legislation as appropriate. Regulatory Preparedness, and Safe-by-Design (SbD) jointly constitute the NanoReg2 Safe Innovation Approach (SIA) for developing innovative products based on nanotechnology. The workshop aimed to gather views and identify current practices in regulatory work on safety of innovative products, tools already in use or needed, and potential difficulties in implementing Regulatory Preparedness in the EU. Presentations addressed the current state of the safety of nanotechnology innovation. The viewpoints included the regulatory framework, the principles behind it and the agencies and authorities enforcing it; nanosafety research projects and their support system (e.g. the current EU Horizon 2020 Framework Programme); national nanosafety initiatives; and the development of tools, such as foresight tools and harmonised test guidelines by the OECD for data generation. The workshop served to generate ideas for achieving Regulatory Preparedness. The participants recognised that while regulators deal with the safety of innovations, only few systematic approaches to this work exist. Some innovative products may reach the market before their safety has been appropriately assessed, as illustrated by RAPEX, the Rapid Exchange of Information System. A continuous and proactive combination of interconnected activities was considered to be required for ensuring Regulatory Preparedness. Thus, anticipation, e.g. horizon scanning, was seen as important, as was communication between regulators, innovators (industry) and other stakeholders. Regulators need to become aware of innovative products under development to ensure that the legislation and methods for safety assessment are available and adequate. Innovators must be aware of regulatory requirements and their likely development. This mutual awareness helps to develop safe products and to avoid delays or other problems in obtaining market approval. Awareness can be achieved through communication, which requires trust, e.g. promoted via "trusted environments" for confidential inquiries and information sharing. Furthermore, regulators need early access to the existing information and data relevant to safety assessment of innovative products to provide timely guidance and advice to Industry as well as to develop strategies for dealing with uncertainty, e.g. by applying the precautionary principle. Regulatory Preparedness was discussed as part of the SIA, and a "road map" of actions was suggested and outlined. The workshop has thus contributed towards acceptance of implementing Regulatory Preparedness for innovation in nanotechnology through the participation of a variety of stakeholders. This paves the way for a better dialogue among stakeholders in a fast economic development cycle, where it is even more important to quickly identify emerging needs for new approaches to regulatory issues for innovationJRC.F.2-Consumer Products Safet

Versailles project on advanced materials and standards (VAMAS) interlaboratory study on measuring the number concentration of colloidal gold nanoparticles

We describe the outcome of a large international interlaboratory study of the measurement of particle number concentration of colloidal nanoparticles, project 10 of the technical working area 34, "Nanoparticle Populations" of the Versailles Project on Advanced Materials and Standards (VAMAS). A total of 50 laboratories delivered results for the number concentration of 30 nm gold colloidal nanoparticles measured using particle tracking analysis (PTA), single particle inductively coupled plasma mass spectrometry (spICP-MS), ultraviolet-visible (UV-Vis) light spectroscopy, centrifugal liquid sedimentation (CLS) and small angle X-ray scattering (SAXS). The study provides quantitative data to evaluate the repeatability of these methods and their reproducibility in the measurement of number concentration of model nanoparticle systems following a common measurement protocol. We find that the population-averaging methods of SAXS, CLS and UV-Vis have high measurement repeatability and reproducibility, with between-labs variability of 2.6%, 11% and 1.4% respectively. However, results may be significantly biased for reasons including inaccurate material properties whose values are used to compute the number concentration. Particle-counting method results are less reproducibile than population-averaging methods, with measured between-labs variability of 68% and 46% for PTA and spICP-MS respectively. This study provides the stakeholder community with important comparative data to underpin measurement reproducibility and method validation for number concentration of nanoparticles

Synthesis, surface modifications and biomedical applications of colloidal gold nanoparticles, towards controlled regulation of angiogenesis in conjunction with photo-thermal therapy

The unique optical properties of colloidal gold nanoparticles: high extinction coefficient and adjustable plasmon band, as well as their excellent biocompatibility (low toxicity and a natural affinity towards thiols and amines) make them very attractive materials for biomedical research. In this project, two types of gold nanoparticles (NPs): spherical and anisotropic were utilised. Spherical NPs predominantly absorb in the visible range of the electromagnetic spectrum, while the plasmon band of anisotropic NPs, e.g. rod-like is shifted into the longer wavelengths. Such variety in the optical signatures of NPs enables their usage in a number of imaging and therapy techniques. Biological activity of NPs is determined by the chemical composition of the organic corona, surrounding the gold core. Appropriate surface capping provides NPs with substantial stability against aggregation (and loss of their properties) in physiological media. Hence, crude colloids were stabilised with a variety of organic molecules, strongly interacting with the gold surface. A selection of functional peptides was incorporated into the organic corona of NPs, in order to achieve desired biological activity of colloids. Functionalised NPs interacted with a certain type of human cells in a selective manner. Such selectivity was obtained by precise recognition between peptides attached to NPs and cell receptors. In this project, human umbilical vein endothelial cells (HUVECs) were studied. These cells build the interior layer of blood vessels in the entire circulatory system and participate in many important physiological processes, as well as pathological developments, e.g. blood vessels formation (angiogenesis). In angiogenesis, several cell receptors are involved. Two of them are: vascular endothelial growth factor receptor type 1 (VEGFR-1) and neuropilin receptor type 1 (NRP-1). Both, VEGFR-1 and NRP-1 were targeted with peptide functionalised NPs. Selectivity of the binding was investigated. Following the binding event, physiological changes in cell metabolism were triggered. The nature and efficiency of these changes were studied, by measuring the expression levels of several angiogenesis related genes, as well as the ability of cells to form capillaries using in vitro angiogenesis assays. HUVECs targeted with anisotropic NPs were illuminated with near-infrared (NIR) laser light. Light at this frequency penetrates human tissue and is absorbed by associated with cells anisotropic NPs. NPs convert the absorbed light into heat, which causes an increase in the local temperature. Increased temperatures result in cell damage and/or stimulate cellular response. Physiological changes in HUVECs upon a moderate heat stimuli were assessed by measuring the expression levels of two genes (ELAM-1 and ELAM-1), which are responsive to the temperature changes. Elevated heat resulted in thermal damage of HUVECs, the degree of which was studied by cell viability assays.In this project, a dual approach towards controlled regulation of HUVECs physiology was demonstrated. Not only regulation of cell metabolism, but the whole angiogenesis process was achieved with peptide functionalised NPs. Targeted with anisotropic NPs cells were illuminated with the NIR laser, which induced the heat shock response in the cell or at the extreme led to death. This work possesses a clear potential and relevance for therapy of various disorders related to insufficient or excessive angiogenesis of endothelial cell