Intracellular uptake, transport, and processing of gold nanostructures

The emerging field of nanomedicine requires better understanding of the interface between nanotechnology and medicine. Better knowledge of the nano-bio interface will lead to better tools for diagnostic imaging and therapy. In this review, recent progress in understanding of how size, shape, and surface properties of nanoparticles (NPs) affect intracellular fate of NPs is discussed. Gold nanostructures are used as a model system in this regard since their physical and chemical properties can be easily manipulated. The NP-uptake is dependent on the physiochemical properties, and once in the cell, most of the NPs are trafficked via an endo-lysosomal path followed by a receptor-mediated endocytosis process at the cell membrane. Within the size range of 2-100 nm, Gold nanoparticles (GNPs) of diameter 50 nm demonstrate the highest uptake. Cellular uptake studies of gold nanorods (GNRs) show that there is a decrease in uptake as the aspect ratio of GNRs increases. Theoretical models support the size- and shape-dependent NP-uptake. The intracellular transport of targeted NPs is faster than untargeted NPs. The surface ligand and charge of NPs play a bigger role in their uptake, transport, and organelle distribution. Exocytosis of NPs is dependent on size and shape as well; however, the trend is different compared to endocytosis. GNPs are now being incorporated into polymer and lipid based NPs to build multifunctional devices. A multifunctional platform based on gold nanostructures, with multimodal imaging, targeting, and therapeutics; hold the possibility of promising directions in medical research.     

Intracellular uptake,transport, and processing of gold nanostructures

Abstract

The emerging field of nanomedicine requires better understanding of the interface between nanotechnology and medicine. Better knowledge of the nano-bio interface will lead to better tools for diagnostic imaging and therapy. In this review, recent progress in understanding of how size, shape, and surface properties of nanoparticles (NPs) affect intracellular fate of NPs is discussed. Gold nanostructures are used as a model system in this regard since their physical and chemical properties can be easily manipulated. The NP-uptake is dependent on the physiochemical properties, and once in the cell, most of the NPs are trafficked via an endo-lysosomal path followed by a receptor-mediated endocytosis process at the cell membrane. Within the size range of 2–100 nm, Gold nanoparticles (GNPs) of diameter 50 nm demonstrate the highest uptake. Cellular uptake studies of gold nanorods (GNRs) show that there is a decrease in uptake as the aspect ratio of GNRs increases. Theoretical models support the size- and shape-dependent NP-uptake. The intracellular transport of targeted NPs is faster than untargeted NPs. The surface ligand and charge of NPs play a bigger role in their uptake, transport, and organelle distribution. Exocytosis of NPs is dependent on size and shape as well; however, the trend is different compared to endocytosis. GNPs are now being incorporated into polymer and lipid based NPs to build multifunctional devices. A multifunctional platform based on gold nanostructures, with multimodal imaging, targeting, and therapeutics; hold the possibility of promising directions in medical research.     

金纳米棒-聚苯胺核壳纳米材料的光学相干层析成像能力研究

对比试剂的使用能够显著提升光学相干层析(OCT)的成像效果。聚苯胺(PANI)是一种有机导电聚合物,在近红外(NIR)区有着很强的光吸收。本文采用PANI对常见的OCT成像对比试剂--金纳米棒(GNRs)进行修饰,合成了PANI/GNRs核壳粒子,并对其OCT成像对比能力进行了研究。PANI/GNRs展现出良好的NIR光吸收特性;同时,PANI对GNRs的包裹也显著提升了金纳米结构的稳定性、降低了GNRs原有的毒性。选用离体猪肝组织作为检测样本,发现纳米材料使用能够显著提升OCT的成像效果。与未修饰的GNRs及PANI粒子相比,PANI/GNRs的OCT成像对比效果明显更好。因此,PANI包裹的GNRs核壳纳米材料有望成为一种低毒性且效果良好的OCT对比试剂用于生物组织成像。     

Investigation of the Plasmonic Interaction of Gold Nanoparticles Toward Plasmonic Photothermal Therapeutics

Plasmonic interaction of nanoparticles located in close proximity, embedded in breast tissue, is simulated for estimating the optical characteristics like optical absorption cross-section, plasmonic wavelength as well as full-width half maxima (FWHM). The computations are done for the monomers, homodimers, and heterodimers of spherical and rod-shaped gold nanoparticles considering various interparticle spacings for gold nanospheres and the interparticle spacing as well as the orientation for gold nanorods (GNRs). The results indicate that for the spherical dimer, with the change in interparticle spacing from 1 to 20 nm, the peak absorption cross-section decreases by 43%. Whereas for the GNRs, the absorption cross-section increases/decreases, within 9–18%, depending on the homodimer or heterodimer configuration. Furthermore, secondary peaks for the absorption cross-section are obtained within wavelengths of 630–940 nm due to antibonding modes for GNR heterodimers. For GNR heterodimer located end-to-end, this secondary peak for the absorption cross-section appears at 780 nm irrespective of interparticle spacing within 1–5 nm. The absorption coefficient is considerably dependent on the configuration and proximity of GNRs located within the tissue. While FWHM is not significantly influenced by GNRs configuration and interparticle spacing. For interparticle spacing from 1 to 20 nm, the plasmonic wavelength shifts by 38 nm for the spherical dimer and by 35–86 nm for various GNR dimers. The findings of this study are useful for plasmonic photothermal therapeutics as the heat generation is governed by the resulting absorption cross-section due to plasmonic coupling of the closely spaced and different orientations of the nanoparticles.

     

In vivo spectroscopic photoacoustic imaging and laser-induced nanoparticle vaporization for anti-HER2 breast cancer

This study reports on the development and application of theragnostic agents targeting the HER2 receptors in breast tumors. The agent was constructed by loading silica-coated gold nanorods (GNRs) and a perfluorohexane liquid into PLGA-PEG nanoparticles, followed by surface conjugation with antibody Herceptin. The particle uptake in human breast cancer MDA-MB-231 (HER2-negative) and BT474 (HER2-positive) cell lines was tested. A proof of principle in vivo study was also performed using a xenograft mouse bilateral tumor model (16 mice, 32 tumors). Photoacoustic imaging was performed using a VevoLAZR device at 720/750/850 nm illuminations and 21 MHz central frequency. The relative concentrations of GNRs in the tumor were quantified using a linear spectral unmixing technique. The therapeutic efficacy of these nanoparticles was evaluated through optical droplet vaporization, and cell damage was confirmed using tissue immunofluorescence and histology. Our results demonstrate the potential of PLGA-GNRs as theragnostic agents for anti-HER2 breast cancer therapy.     

Rapid and specific influenza virus detection by functionalized magnetic nanoparticles and mass spectrometry

Background

When a fluorophore is placed in the vicinity of a metal nanoparticle possessing a strong plasmon field, its fluorescence emission may change extensively. Our study is to better understand this phenomenon and predict the extent of quenching and/or enhancement of fluorescence, to beneficially utilize it in molecular sensing/imaging.

Results

Plasmon field intensities on/around gold nanoparticles (GNPs) with various diameters were theoretically computed with respect to the distance from the GNP surface. The field intensity decreased rapidly with the distance from the surface and the rate of decrease was greater for the particle with a smaller diameter. Using the plasmon field strength obtained, the level of fluorescence alternation by the field was theoretically estimated. For experimental studies, 10 nm GNPs were coated with polymer layer(s) of known thicknesses. Cypate, a near infrared fluorophore, was placed on the outermost layer of the polymer coated GNPs, artificially separated from the GNP at known distances, and its fluorescence levels were observed. The fluorescence of Cypate on the particle surface was quenched almost completely and, at approximately 5 nm from the surface, it was enhanced ~17 times. The level decreased thereafter. Theoretically computed fluorescence levels of the Cypate placed at various distances from a 10 nm GNP were compared with the experimental data. The trend of the resulting fluorescence was similar. The experimental results, however, showed greater enhancement than the theoretical estimates, in general. The distance from the GNP surface that showed the maximum enhancement in the experiment was greater than the one theoretically predicted, probably due to the difference in the two systems.

Conclusions

Factors affecting the fluorescence of a fluorophore placed near a GNP are the GNP size, coating material on GNP, wavelengths of the incident light and emitted light and intrinsic quantum yield of the fluorophore. Experimentally, we were able to quench and enhance the fluorescence of Cypate, by changing the distance between the fluorophore and GNP. This ability of artificially controlling fluorescence can be beneficially used in developing contrast agents for highly sensitive and specific optical sensing and imaging.     

The Tunable and Well-Controlled Surface Plasmon Resonances of Au Hollow Nanostructures by a Chemical Route

Au plasmonic hollow spherical nanostructures were synthesized by electrochemical reduction (GRR, the Galvanic Replacement Reaction) using Ag nanoparticles as templates. From UV-visible absorption spectroscopy, it was found that the surface plasmon resonance (SPR) of gold hollow spherical nanostructures first showed red shift and then blue shift. However, further addition of gold precursor (HAuCl₄) resulted into a red shift of SPR peak. The morphological changes from Ag nanoparticles to Au hollow nanostructures were assessed by transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDX)analysis. The Mie Scattering theory based simulations of SPR of Au hollow nanostructures were performed which are in good agreement with the experimental observations. Based on the experimental observations and theoretical calculations, a complete growth mechanism for Au hollow nanostructures is proposed.     

Gold(III) Reduction by the Rhizobacterium Azospirillum brasilense with the Formation of Gold Nanoparticles

For the soil nitrogen-fixing bacterium Azospirillum brasilense, the ability to reduce [AuCl₄]^? and to form gold nanoparticles (GNPs) has been demonstrated, with the appearance of a mauve tint of the culture. To validate the shapes and chemical nature of nanoparticles, transmission electron microscopy (TEM) and X-ray fluorescence analysis were used. For the widely studied agriculturally important wild-type strains A. brasilense Sp7 and Sp245, GNPs formed after 10 days of incubation of cell biomass with 0.25 mM [AuCl₄]^? were shown (using TEM) to be mainly of spherical form (5 to 20 nm in diameter), with rare occasional triangles. In the course of cultivation with [AuCl₄]^?, after 5 days, a mauve tint was already visible for cells of strain Sp245.5, after 6 days for Sp245 and after 10 days for Sp7. Thus, for the mutant strain Sp245.5 (which has significant differences in the structure and composition of cell-surface polysaccharides as compared with Sp245), a more rapid formation of GNPs was observed. Moreover, their TEM images (also obtained after 10 days) showed different shapes: nano-sized spheres, triangles, hexagons and rods, as well as larger round-shaped flower-like nanoparticles about 100 nm in size. Since by the time of GNP formation in our experiments the cells were found to be already not viable, this confirms the dominating role of cell surface structure and chemical composition in shaping the GNPs formed in the course of [AuCl₄]^? reduction to Au⁰. This finding may be useful for understanding the natural biogeochemical mechanisms of gold reduction and formation of GNPs involving microorganisms. The data obtained may also help in developing protocols for environmentally friendly synthesis of nanoparticles and possible use of bacterial cells with modified surface structure and composition for their fabrication.     

Biological synthesis of gold nanoparticles using Magnolia kobus and Diopyros kaki leaf extracts

Leaf extracts of two plants, Magnolia kobus and Diopyros kaki, were used for ecofriendly extracellular synthesis of metallic gold nanoparticles. Stable gold nanoparticles were formed by treating an aqueous HAuCl₄ solution using the plant leaf extracts as reducing agents. UV–visible spectroscopy was used for quantification of gold nanoparticle synthesis. Only a few minutes were required for >90% conversion to gold nanoparticles at a reaction temperature of 95 °C, suggesting reaction rates higher or comparable to those of nanoparticle synthesis by chemical methods. The synthesized gold nanoparticles were characterized with inductively coupled plasma spectrometry (ICP), energy-dispersive X-ray spectroscopy (EDS), scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), Fourier-transform infrared spectroscopy (FTIR), and particle analysis using a particle analyzer. SEM and TEM images showed that a mixture of plate (triangles, pentagons, and hexagons) and spherical structures (size, 5–300 nm) were formed at lower temperatures and leaf broth concentrations, while smaller spherical shapes were obtained at higher temperatures and leaf broth concentrations.     

Green biochemistry approach for synthesis of silver and gold nanoparticles using <Emphasis Type="Italic">Ficus racemosa</Emphasis> latex and their pH-dependent binding study with different amino acids using UV/Vis absorption spectroscopy

Simple and eco-friendly biosynthesis approach was developed to synthesize silver nanoparticles (SNPs) and gold nanoparticles (GNPs) using Ficus racemosa latex as reducing agent. The presence of sunlight is utilized with latex and achieved the nanoparticles whose average size was in the range of 50–120 nm for SNPs and 20–50 nm for GNPs. The synthesized nanoparticles were characterized by UV/Visible absorption spectroscopy, X-ray diffraction, and field emission—scanning electron microscopy techniques toget understand the obtained nanoparticles. The pH-dependent binding studies of SNPs and GNPs with four amino acids, namely l-lysine, l-arginine, l-glutamine and glycin have been reported.     

Bio-Synthesis of Gold Nanoparticles by <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> and Assessment of Their Conjugation Possibility with Two Types of β-Lactam Antibiotics without Any Additional Linkers

In the present study, biosynthesis of gold nanoparticles (GNPs) by Fusarium oxysporum was carried out and their conjugation possibility with two β-lactam antibiotics was evaluated. F. oxysporum was cultured and the fungal culture supernatant was subjected to the 1 mmol final concentration of chloroauric acid solution. The produced GNPs were analyzed using visible spectrophotometer, X-ray diffraction analysis (XRD) and transmission electron microscope (TEM). After the purification of GNPs, they were subjected to penicillin G and ceftriaxone without any additional linkers. Finally, the mixture was analyzed using visible spectrophotometer, Fourier transform infrared spectroscopy (FTIR) and TEM and subjected to antibacterial activity test using the well diffusion method. Results confirmed the presence of GNPs in the F. oxysporum culture supernatant after the addition of chloroauric acid. TEM results showed that GNPs were spherical and amorphous with sizes around 10–25 nm and XRD confirmed the presence of GNPs in the fungal culture supernatant. After the incubation of GNPs with the antibiotics, FTIR results demonstrated the successful linking of GNPs with the corresponded antibiotics and TEM images showed that GNPs sizes were bigger than the pure ones (around 50–100 nm). Finally the antibacterial activity test indicated that absolutely, the antibacterial properties of the GNPs-β-lactam antibiotics was lowered or not changed in contrast to the pure antibiotics. The present study showed that GNPs had high tendency of conjugation with antibiotics but unlike the previous researches, linking of the antibiotics to GNPs always cannot improve their antibacterial activity based on the antibiotics that were used. The high conjugation affinity of GNPs made them a good candidate as detoxification agent in diverse areas of medicine or environmental sciences.     

Oligonucleotide-coated metallic nanoparticles as a flexible platform for molecular imaging agents

Targeted metallic nanoparticles have shown promise as contrast agents for molecular imaging. To obtain molecular specificity, the nanoparticle surface must be appropriately functionalized with probe molecules that will bind to biomarkers of interest. The aim of this study was to develop and characterize a flexible approach to generate molecular imaging agents based on gold nanoparticles conjugated to a diverse range of probe molecules. We present two complementary oligonucleotide-based approaches to develop gold nanoparticle contrast agents which can be functionalized with a variety of biomolecules ranging from small molecules, to peptides, to antibodies. The size, biocompatibility, and protein concentration per nanoparticle are characterized for the two oligonucleotide-based approaches; the results are compared to contrast agents prepared using adsorption of proteins on gold nanoparticles by electrostatic interaction. Contrast agents prepared from oligonucleotide-functionalized nanoparticles are significantly smaller in size and more stable than contrast agents prepared by adsorption of proteins on gold nanoparticles. We demonstrate the flexibility of the oligonucleotide-based approach by preparing contrast agents conjugated to folate, EGF peptide, and anti-EGFR antibodies. Reflectance images of cancer cell lines labeled with functionalized contrast agents show significantly increased image contrast which is specific for the target biomarker. To demonstrate the modularity of this new bioconjugation approach, we use it to conjugate both fluorophore and anti-EGFR antibodies to metal nanoparticles, yielding a contrast agent which can be probed with multiple imaging modalities. This novel bioconjugation approach can be used to prepare contrast agents targeted with biomolecules that span a diverse range of sizes; at the same time, the bioconjugation method can be adapted to develop multimodal contrast agents for molecular imaging without changing the coating design or material.     

Biosynthesis of extracellular and intracellular gold nanoparticles by Aspergillus fumigatus and A. flavus

Green chemistry is a boon for the development of safe, stable and ecofriendly nanostructures using biological tools. The present study was carried out to explore the potential of selected fungal strains for biosynthesis of intra- and extracellular gold nanostructures. Out of the seven cultures, two fungal strains (SBS-3 and SBS-7) were selected on the basis of development of dark pink colour in cell free supernatant and fungal beads, respectively indicative of extra- and intracellular gold nanoparticles production. Both biomass associated and cell free gold nanoparticles were characterized using X-ray diffractogram (XRD) analysis and transmission electron microscopy (TEM). XRD analysis confirmed crystalline, face-centered cubic lattice of metallic gold nanoparticles along with average crystallite size. A marginal difference in average crystallite size of extracellular (17.76 nm) and intracellular (26 and 22 nm) Au-nanostructures was observed using Scherrer equation. In TEM, a variety of shapes (triangles, spherical, hexagonal) were observed in both extra- and intracellular nanoparticles. 18S rRNA gene sequence analysis by multiple sequence alignment (BLAST) indicated 99 % homology of SBS-3 to Aspergillus fumigatus with 99 % alignment coverage and 98 % homology of SBS-7 to Aspergillus flavus with 98 % alignment coverage respectively. Native-PAGE and activity staining further confirmed enzyme linked synthesis of gold nanoparticles.     

Targeted radiotherapy enhancement during electronic brachytherapy of accelerated partial breast irradiation (APBI) using controlled release of gold nanoparticles

Several studies have demonstrated low rates of local recurrence with brachytherapy-based accelerated partial breast irradiation (APBI). However, long-term outcomes on toxicity (e.g. telangiectasia) and cosmesis remain a major concern. The purpose of this study is to investigate the dosimetric feasibility of using targeted non-toxic radiosensitizing gold nanoparticles (GNPs) for localized dose enhancement to the planning target volume (PTV) during electronic brachytherapy APBI while reducing normal tissue toxicity. We propose to incorporate GNPs into a micrometer-thick polymer film on the surface of routinely used lumpectomy balloon applicators and provide subsequent treatment using a 50 kVp Xoft device. An experimentally determined diffusion coefficient was used to determine space-time customizable distribution of GNPs for feasible in-vivo concentrations of 7 mg/g and 43 mg/g. An analytical approach from previously published work was employed to estimate the dose enhancement due to GNPs as a function of distance up to 1 cm from the lumpectomy cavity surface. Clinically significant dose enhancement values of at least 1.2, due to 2 nm GNPs, were found at 1 cm away from the lumpectomy cavity wall when using electronic brachytherapy APBI. Higher customizable dose enhancement was also achieved at other distances as a function of nanoparticle size. Our preliminary results suggest that significant dose enhancement can be achieved to residual tumor cells targeted with GNPs during APBI with electronic brachytherapy.     

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.     

Intercomparison of dose enhancement ratio and secondary electron spectra for gold nanoparticles irradiated by X-rays calculated using multiple Monte Carlo simulation codes

PurposeTargeted radiation therapy has seen an increased interest in the past decade. In vitro and in vivo experiments showed enhanced radiation doses due to gold nanoparticles (GNPs) to tumors in mice and demonstrated a high potential for clinical application. However, finding a functionalized molecular formulation for actively targeting GNPs in tumor cells is challenging. Furthermore, the enhanced energy deposition by secondary electrons around GNPs, particularly by short-ranged Auger electrons is difficult to measure. Computational models, such as Monte Carlo (MC) radiation transport codes, have been used to estimate the physical quantities and effects of GNPs. However, as these codes differ from one to another, the reliability of physical and dosimetric quantities needs to be established at cellular and molecular levels, so that the subsequent biological effects can be assessed quantitatively.MethodsIn this work, irradiation of single GNPs of 50 nm and 100 nm diameter by X-ray spectra generated by 50 and 100 peak kilovoltages was simulated for a defined geometry setup, by applying multiple MC codes in the EURADOS framework.ResultsThe mean dose enhancement ratio of the first 10 nm-thick water shell around a 100 nm GNP ranges from 400 for 100 kVp X-rays to 600 for 50 kVp X-rays with large uncertainty factors up to 2.3.ConclusionsIt is concluded that the absolute dose enhancement effects have large uncertainties and need an inter-code intercomparison for a high quality assurance; relative properties may be a better measure until more experimental data is available to constrain the models.     

An in-silico method to predict and quantify the effect of gold nanoparticles in X-ray imaging

PurposeOver the last few years studies are conducted, highlighting the feasibility of Gold Nanoparticles (GNPs) to be used in clinical CT imaging and as an efficient contrast agent for cancer research. After ensuring that GNPs formulations are appropriate for in vivo or clinical use, the next step is to determine the parameters for an X-ray system’s optimal contrast for applications and to extract quantitative information. There is currently a gap and need to exploit new X-ray imaging protocols and processing algorithms, through specific models avoiding trial-and-error procedures and provide an imaging prognosis tool. Such a model can be used to confirm the accumulation of GNPs in target organs before radiotherapy treatments with a system easily available in hospitals, as low energy X-rays.MethodsIn this study a complete, easy-to-use, simulation platform is designed and built, where simple parameters, as the X-ray’s specifications and experimentally defined biodistributions of specific GNPs are imported. The induced contrast and images can be exported, and accurate quantification can be performed. This platform is based on the GATE Monte Carlo simulation toolkit, based on the GEANT4 toolkit and the MOBY phantom, a realistic 4D digital mouse.ResultsWe have validated this simulation platform to predict the contrast induction and minimum detectable concentration of GNPs on any given X-ray system. The study was applied to preclinical studies but is also expandable to clinical studies.ConclusionsAccording to our knowledge, no other such validated simulation model currently exists, and this model could help radiology imaging with GNPs to be truly deployed.     

Inorganic nanoparticle-based contrast agents for molecular imaging

Inorganic nanoparticles (NPs) including semiconductor quantum dots (QDs), iron oxide NPs and gold NPs have been developed as contrast agents for diagnostics by molecular imaging. Compared with traditional contrast agents, NPs offer several advantages: their optical and magnetic properties can be tailored by engineering the composition, structure, size and shape; their surfaces can be modified with ligands to target specific biomarkers of disease; the contrast enhancement provided can be equivalent to millions of molecular counterparts; and they can be integrated with a combination of different functions for multimodal imaging. Here, we review recent advances in the development of contrast agents based on inorganic NPs for molecular imaging, and also touch on contrast enhancement, surface modification, tissue targeting, clearance and toxicity. As research efforts intensify, contrast agents based on inorganic NPs that are highly sensitive, target-specific and safe to use are expected to enter clinical applications in the near future.     

Bioaccumulation and toxicity of gold nanoparticles after repeated administration in mice

Gold nanoparticles (GNPs) offer a great promise in biomedicine. Currently, there is no data available regarding the accumulation of nanoparticles in vivo after repeated administration. The purpose of the present study was to evaluate the bioaccumulation and toxic effects of different doses (40, 200, and 400 μg/kg/day) of 12.5 nm GNPs upon intraperitoneal administration in mice every day for 8 days.The gold levels in blood did not increase with the dose administered, whereas in all the organs examined there was a proportional increase on gold, indicating efficient tissue uptake. Although brain was the organ containing the lowest quantity of injected GNPs, our data suggest that GNPs are able to cross the blood-brain barrier and accumulate in the neural tissue. Importantly, no evidence of toxicity was observed in any of the diverse studies performed, including survival, behavior, animal weight, organ morphology, blood biochemistry and tissue histology. The results indicate that tissue accumulation pattern of GNPs depend on the doses administered and the accumulation of the particles does not produce sub-acute physiological damage.     

Cellular Uptake of Gold Nanoparticles and Their Behavior as Labels for Localization Microscopy

Gold nanoparticles (GNPs) enhance the damaging absorbance effects of high-energy photons in radiation therapy by increasing the emission of Auger-photoelectrons in the nm-μm range. It has been shown that the incorporation of GNPs has a significant effect on radiosensitivity of cells and their dose-dependent clonogenic survival. One major characteristic of GNPs is also their diameter-dependent cellular uptake and retention. In this article, we show by means of an established embodiment of localization microscopy, spectral position determination microscopy (SPDM), that imaging with nanometer resolution and systematic counting of GNPs becomes feasible, because optical absorption and plasmon resonance effects result in optical blinking of GNPs at a size-dependent wavelength. To quantify cellular uptake and retention or release, SPDM with GNPs that have diameters of 10 and 25 nm was performed after 2 h and after 18 h. The uptake of the GNPs in HeLa cells was either achieved via incubation or transfection via DNA labeling. On average, the uptake by incubation after 2 h was approximately double for 10 nm GNPs as compared to 25 nm GNPs. In contrast, the uptake of 25 nm GNPs by transfection was approximately four times higher after 2 h. The spectral characteristics of the fluorescence of the GNPs seem to be environment-dependent. In contrast to fluorescent dyes that show blinking characteristics due to reversible photobleaching, the blinking of GNPs seems to be stable for long periods of time, and this facilitates their use as an appropriate dye analog for SPDM imaging.