Use of Coated Silver Nanoparticles to Understand the Relationship of Particle Dissolution and Bioavailability to Cell and Lung Toxicological Potential

Since more than 30% of consumer products that include engineered nanomaterials contain nano‐Ag, the safety of this material is of considerable public concern. In this study, Ag nanoparticles (NPs) are used to demonstrate that 20 nm polyvinylpyrrolidone (PVP or P) and citrate (C)‐coated Ag NPs induce more cellular toxicity and oxidative stress than larger (110 nm) particles due to a higher rate of dissolution and Ag bioavailability. Moreover, there is also a higher propensity for citrate 20 nm (C20) nanoparticles to generate acute neutrophilic inflammation in the lung and to produce chemokines compared to C110. P110 has less cytotoxic effects than C110, likely due to the ability of PVP to complex released Ag⁺. In contrast to the more intense acute pulmonary effects of C20, C110 induces mild pulmonary fibrosis at day 21, likely as a result of slow but persistent Ag⁺ release leading to a sub‐chronic injury response. Interestingly, the released metallic Ag is incorporated into the collagen fibers depositing around airways and the lung interstitium. Taken together, these results demonstrate that size and surface coating affect the cellular toxicity of Ag NPs as well as their acute versus sub‐chronic lung injury potential.     

Quantitative Evaluation of Cellular Uptake and Trafficking of Plain and Polyethylene Glycol‐Coated Gold Nanoparticles

This study addresses the cellular uptake and intracellular trafficking of 15‐nm gold nanoparticles (NPs), either plain (i.e., stabilized with citrate) or coated with polyethylene glycol (PEG), exposed to human alveolar epithelial cells (A549) at the air–liquid interface for 1, 4, and 24 h. Quantitative analysis by stereology on transmission electron microscopy images reveals a significant, nonrandom intracellular distribution for both NP types. No particles are observed in the nucleus, mitochondria, endoplasmic reticulum, or golgi. The cytosol is not a preferred cellular compartment for both NP types, although significantly more PEG‐coated than citrate‐stabilized NPs are present there. The preferred particle localizations are vesicles of different sizes (<150, 150–1000, >1000 nm). This is observed for both NP types and indicates a predominant uptake by endocytosis. Subsequent inhibition of caveolin‐ and clathrin‐mediated endocytosis by methyl‐β‐cyclodextrin (MβCD) results in a significant reduction of intracellular NPs. The inhibition, however, is more pronounced for PEG‐coated than citrate‐stabilized NPs. The latter are mostly found in larger vesicles; therefore, they are potentially taken up by macropinocytosis, which is not inhibited by MβCD. With prolonged exposure times, both NPs are preferentially localized in larger‐sized intracellular vesicles such as lysosomes, thus indicating intracellular particle trafficking. This quantitative evaluation reveals that NP surface coatings modulate endocytotic uptake pathways and cellular NP trafficking. Other nonendocytotic entry mechanisms are found to be involved as well, as indicated by localization of a minority of PEG‐coated NPs in the cytosol.     

Protein Corona Mediated Uptake and Cytotoxicity of Silver Nanoparticles in Mouse Embryonic Fibroblast

Medical applications of nanoparticles (NPs) require understanding of their interactions with living systems in order to control their physiological response, such as cellular uptake and cytotoxicity. When NPs are exposed to biological fluids, the adsorption of extracellular proteins on the surface of NPs, creating the so‐called protein corona, can critically affect their interactions with cells. Here, the effect of surface coating of silver nanoparticles (AgNPs) on the adsorption of serum proteins (SPs) and its consequence on cellular uptake and cytotoxicity in mouse embryonic fibroblasts are shown. In particular, citrate‐capped AgNPs are internalized by cells and show a time‐ and dose‐dependent toxicity, while the passivation of the NP surface with an oligo(ethylene glycol) (OEG)‐alkanethiol drastically reduces their uptake and cytotoxicity. The exposure to growth media containing SPs reveals that citrate‐capped AgNPs are promptly coated and stabilized by proteins, while the AgNPs resulting from capping with the OEG‐alkanethiol are more resistant to adsorption of proteins onto their surface. Using NIH‐3T3 cultured in serum‐free, the key role of the adsorption of SPs onto surface of NPs is shown as only AgNPs with a preformed protein corona can be internalized by the cells and, consequently, carry out their inherent cytotoxic activity.     

Implications of silver nanoparticle induced cell apoptosis for in vitro gene therapy

The impact of manufactured nanomaterials on human health and the environment is a major concern for commercial use of nanotechnology based products. A judicious choice of selective usage, lower nanomaterial concentration and use in combination with conventional therapeutic materials may provide the best solution. For example, silver nanoparticles (Ag NPs) are known to be bactericidal and also cytotoxic to mammalian cells. Herein, we investigate the molecular mechanism of Ag NP mediated cytotoxicity in both cancer and non-cancer cells and find that optimum particle concentration leads to programmed cell death in vitro. Also, the benefit of the cytotoxic effects of Ag NPs was tested for therapeutic use in conjunction with conventional gene therapy. The synergistic effect of Ag NPs on the uracil phosphoribosyltransferase expression system sensitized the cells more towards treatment with the drug 5-fluorouracil. Induction of the apoptotic pathway makes Ag NPs a representative of a new chemosensitization strategy for future application in gene therapy.     

Tuning Cellular Response to Nanoparticles via Surface Chemistry and Aggregation

The aggregation of gold nanoparticles (Au NPs) in cell media is a common phenomenon that can influence NP‐cell interactions. Here, we control Au NP aggregation in cell media and study the impact of Au NP aggregation on human dermal fibroblast (HDF) cells. By first adding Au NPs to fetal bovine serum (FBS) and then subsequently to a buffer, aggregation can be avoided. Aggregation of Au NPs also can be avoided by coating Au NPs with other biomolecules such as lipids. The aggregation state of the Au NPs influences cellular toxicity and Au NP uptake: non‐aggregated cationic Au NPs are four‐fold less toxic to HDF cells than aggregated cationic Au NPs, and the uptake of non‐aggregated anionic citrate Au NPs is three orders of magnitude less than that of aggregated citrate Au NPs. Upon uptake of Au NPs, cellular F‐actin fiber formation is disrupted and actin dots are predominant. When lipid‐coated Au NPs are doped with a fluorescent lipid (F‐lipid) and incubated with HDF cells, the fluorescence from the F‐lipid was found throughout the cell, showing that lipids can dissociate from the Au NP surface upon entering the cell.     

Ultrafine Silver Nanoparticles Embedded in Cyclodextrin Metal‐Organic Frameworks with GRGDS Functionalization to Promote Antibacterial and Wound Healing Application

The challenge of bacterial infection increases the risk of mortality and morbidity in acute and chronic wound healing. Silver nanoparticles (Ag NPs) are a promising new version of conventional antibacterial nanosystem to fight against the bacterial resistance in concern of the drug discovery void. However, there are several challenges in controlling the size and colloidal stability of Ag NPs, which readily aggregate or coalesce in both solid and aqueous state. In this study, a template‐guided synthesis of ultrafine Ag NPs of around 2 nm using water‐soluble and biocompatible γ‐cyclodextrin metal‐organic frameworks (CD‐MOFs) is reported. The CD‐MOF based synthetic strategy integrates AgNO₃ reduction and Ag NPs immobilization in one pot achieving dual functions of reduced particle size and enhanced stability. Meanwhile, the synthesized Ag NPs are easily dispersible in aqueous media and exhibit effective bacterial inhibition. The surface modification of cross‐linked CD‐MOF particles with GRGDS peptide boosts the hemostatic effect that further enhances wound healing in synergy with the antibacterial effect. Hence, the strategy of ultrafine Ag NPs synthesis and immobilization in CD‐MOFs together with GRGDS modification holds promising potential for the rational design of effective wound healing devices.     

Uptake Kinetics and Nanotoxicity of Silica Nanoparticles Are Cell Type Dependent

In this study, it is shown that the cytotoxic response of cells as well as the uptake kinetics of nanoparticles (NPs) is cell type dependent. We use silica NPs with a diameter of 310 nm labeled with perylene dye and 304 nm unlabeled particles to evaluate cell type‐dependent uptake and cytotoxicity on human vascular endothelial cells (HUVEC) and cancer cells derived from the cervix carcinoma (HeLa). Besides their size, the particles are characterized concerning homogeneity of the labeling and their zeta potential. The cellular uptake of the labeled NPs is quantified by imaging the cells via confocal microscopy in a time‐dependent manner, with subsequent image analysis via a custom‐made and freely available digital method, Particle_in_Cell‐3D. We find that within the first 4 h of interaction, the uptake of silica NPs into the cytoplasm is up to 10 times more efficient in HUVEC than in HeLa cells. Interestingly, after 10 or 24 h of interaction, the number of intracellular particles for HeLa cells by far surpasses the one for HUVEC. Inhibitor studies show that these endothelial cells internalize 310 nm SiO₂ NPs via the clathrin‐dependent pathway. Remarkably, the differences in the amount of taken up NPs are not directly reflected by the metabolic activity and membrane integrity of the individual cell types. Interaction with NPs leads to a concentration‐dependent decrease in mitochondrial activity and an increase in membrane leakage for HUVEC, whereas HeLa cells show only a reduced mitochondrial activity and no membrane leakage. In addition, silica NPs lead to HUVEC cell death while HeLa cells survive. These findings indicate that HUVEC are more sensitive than HeLa cells upon silica NP exposure.     

Cetyltrimethylammonium bromide-modified spherical and cube-like gold nanoparticles as extrinsic Raman labels in surface-enhanced Raman spectroscopy based heterogeneous immunoassays

This paper reports on the characterization and preliminary comparison of gold nanoparticles of differing surface modification and shape when used as extrinsic Raman labels (ERLs) in high-sensitivity heterogeneous immunoassays based on surface enhanced Raman scattering (SERS). ERLs are gold nanoparticles coated with an adlayer of an intrinsically strong Raman scatterer, followed by a coating of a molecular recognition element (e.g., antibody). Three types of ERLs, all with a nominal size of approximately 30 nm, were fabricated by using spherical citrate-capped gold nanoparticles (sp-cit-Au NPs), spherical CTAB-capped gold nanoparticles (sp-CTAB-Au NPs), or cube-like CTAB-capped gold nanoparticles (cu-CTAB-Au NPs) as cores. The performance of these particles was assessed via a sandwich immunoassay for human IgG in phosphate buffered saline. The ERLs fabricated with sp-CTAB-Au NPs as cores proved to be more than 50 times more sensitive than those with sp-cit-Au NPs as cores; the same comparison showed that the ERLs with cu-CTAB-Au NPs as cores were close to 200 times more sensitive. Coupled with small differences in levels of nonspecific adsorption, these sensitivities translated to a limit of detection (LOD) of 94, 2.3, and 0.28 ng/mL, respectively, for the detection of human IgG in the case of sp-cit-Au NPs, sp-CTAB-Au NPs, and cu-CTAB-Au NPs. The LOD of the cu-CTAB-Au NPs is therefore approximately 340 times below that for the sp-cit-Au NPs. Potential applications of these labels to bioassays are briefly discussed.     

Induction of apoptosis in cancer cells at low silver nanoparticle concentrations using chitosan nanocarrier

We report the development of a chitosan nanocarrier (NC)-based delivery of silver nanoparticles (Ag NPs) to mammalian cells for induction of apoptosis at very low concentrations of the NPs. The cytotoxic efficacy of the Ag NP-nanocarrier (Ag-CS NC) system in human colon cancer cells (HT 29) was examined by morphological analyses and biochemical assays. Cell viability assay demonstrated that the concentration of Ag NPs required to reduce the viability of HT 29 cells by 50% was 0.33 μg mL(-1), much less than in previously reported data. The efficient induction of apoptosis by Ag-CS NCs was confirmed by flow cytometry. Additionally, the characteristic nuclear and morphological changes during apoptotic cell death were investigated by fluorescence and scanning electron microscopy (SEM), respectively. The involvement of mitochondrial pathway of cell death in the Ag-CS NCs induced apoptosis was evident from the depolarization of mitochondrial membrane potential (ΔΨ(m)). Real time quantitative RT-PCR analysis demonstrated the up-regulation of caspase 3 expression which was further reflected in the formation of oligo-nucleosomal DNA "ladders" in Ag-CS NCs treated cells, indicating the important role of caspases in the present apoptotic process. The increased production of intracellular ROS due to Ag-CS NCs treatment indicated that the oxidative stress could augment the induction of apoptosis in HT 29 cells in addition to classical caspase signaling pathway. The use of significantly low concentration of Ag NPs impregnated in chitosan nanocarrier is a much superior approach in comparison to the use of free Ag NPs in cancer therapy.     

Amorphous Silica Nanoparticles Promote Monocyte Adhesion to Human Endothelial Cells: Size‐Dependent Effect

There is evidence that nanoparticles can induce endothelial dysfunction. Here, the effect of monodisperse amorphous silica nanoparticles (SiO₂‐NPs) of different diameters on endothelial cells function is examined. Human endothelial cell line (EA.hy926) or primary human pulmonary artery endothelial cells (hPAEC) are seeded in inserts introduced or not above triple cell co‐cultures (pneumocytes, macrophages, and mast cells). Endothelial cells are incubated with SiO₂‐NPs at non‐cytotoxic concentrations for 12 h. A significant increase (up to 2‐fold) in human monocytes adhesion to endothelial cells is observed for 18 and 54 nm particles. Exposure to SiO₂‐NPs induces protein expression of adhesion molecules (ICAM‐1 and VCAM‐1) as well as significant up‐regulation in mRNA expression of ICAM‐1 in both endothelial cell types. Experiments performed with fluorescent‐labelled monodisperse amorphous SiO₂‐NPs of similar size evidence nanoparticle uptake into the cytoplasm of endothelial cells. It is concluded that exposure of human endothelial cells to amorphous silica nanoparticles enhances their adhesive properties. This process is modified by the size of the nanoparticle and the presence of other co‐cultured cells.     

In-vivo and in-vitro biocompatibility evaluations of silver nanoparticles with antimicrobial activity

In this study, the biocompatibility and antimicrobial activity of silver nanoparticles (Ag NPs) were evaluated in vitro and in vivo. The cytotoxicity of Ag NPs (average diameter: 2-5 nm) against CHO-K1 cells was determined via WST-8 assay, and their genotoxicity was examined via Salmonella typhimurium reverse mutation assay (Ames test). The acute toxicity and intracutaneous reactivity of Ag NPs were evaluated using animal models of mice and rabbits, respectively. The antibacterial effects of Ag NPs on the Gram (-) bacterial strains of Escherichia coli ATCC 8739 and Pseudomonas aeruginosa ATCC 9027 and on the Gram (+) bacterial strains of Staphylococcus aureus ATCC 6538p and Bacillus subtilius ATCC 6633 were determined by measuring the minimum inhibitory concentrations. The Ag NPs were highly cytotoxic to the L-929 cells at over 2 ppm but were non-cytotoxic at lower than 1 ppm. Moreover, the Ag NPs at 1 ppm or lower did not show genotoxicity, acute toxicity and intracutaneous reactivity. It was also found that the Ag NPs exerted effective antimicrobial activities on both the Gram (-) and (+) bacterial strains within the range from 0.06 to 0.98 ppm for 50% MIC.     

Incorporated Organic Modified Ag Nanoparticles in Ormocer

Ag nanoparticles coated trisodium citrate were incorporated in ormocer by sol-gel method. The doping concentration of Ag in ormocer is about 1.0% in weight. The HRTEM demonstrated that the particles disperse in ormocer, and the size of Ag nanoparticles is 5-10 nm. The absorption band of Ag nanoparticle at 410 nm was observed.     

Effect of the surface texture and crystallinity of ZnO nanoparticles on their toxicity

We have investigated the correlation between the structural properties of ZnO nanoparticles (NPs) and their toxicity to mesenchymal stem cells (C2C12 cell line) and macrophage-derived cells (RAW 264.7 cell line). Nanopowders of grain size ranging between 5 nm and 50 nm were prepared by chemical route. Their structural properties were characterized extensively by X-ray Diffraction (XRD) and High Resolution Transmission Electron Microscopy (HRTEM). The XRD spectra showed that 50 nm sized NPs are well crystallized and present a preferential orientation along the direction normal to the (001) plane while the HREM observations revealed that most of the large size (50 nm) crystallized nanoparticles have polygonal shape which is consistent with a texture of along [001] direction. The toxicity tests showed that [001] large textured NPs have higher toxicity to inflammatory cells than nanoparticles of low crystallinity and much smaller size (5 nm). In addition, NPs have cytotoxic effects on inflammatory cells at concentration as low as 0.05 mM while ten times higher concentrations did not have significant cytotoxic effects on cells representing mesenchymal tissues. These observations are explained by the enhanced generation of Reactive Oxygen Species (ROS) at the (0001) polar surface of ZnO NP. These results provide a direct evidence of the correlation between the toxicity and the surface texture of the oxide nanoparticles. Similar correlation has been reported for the photocatalytic properties of ZnO nanoparticles.     

Mechanistic Investigation of the Biological Effects of SiO2, TiO2, and ZnO Nanoparticles on Intestinal Cells

Silicon dioxide (SiO₂), titanium dioxide (TiO₂), and zinc oxide (ZnO) are currently among the most widely used nanoparticles (NPs) in the food industry. This could potentially lead to unintended exposure of the gastrointestinal tract to these NPs. This study aims to investigate the potential side‐effects of these food‐borne NPs on intestinal cells and to mechanistically understand the observed biological responses. Among the panel of tested NPs, ZnO NPs are the most toxic. Consistently in all three tested intestinal cell models, ZnO NPs invoke the most inflammatory responses from the cells and induce the highest intracellular production of reactive oxygen species (ROS). The elevated ROS levels induce significant damage to the DNA of the cells, resulting in cell‐cycle arrest and subsequently cell death. In contrast, both SiO₂ and TiO₂ NPs elicit minimum biological responses from the intestinal cells. Overall, the study showcases the varying capability of the food‐borne NPs to induce a cellular response in the intestinal cells. In addition to physicochemical differences in the NPs, the genetic landscape of the intestinal cell models governs the toxicology profile of these food‐borne NPs.     

Mesoporous Silica Supported Silver–Bismuth Nanoparticles as Photothermal Agents for Skin Infection Synergistic Antibacterial Therapy

The emergence of multidrug resistant bacteria has resulted in plenty of stubborn nosocomial infections and severely threatens human health. Developing novel bactericide and therapeutic strategy is urgently needed. Herein, mesoporous silica supported silver–bismuth nanoparticles (Ag‐Bi@SiO₂ NPs) are constructed for synergistic antibacterial therapy. In vitro experiments indicate that the hyperthermia originating from Bi NPs can disrupt cell integrity and accelerate the Ag ions release, further exhibiting an excellent antibacterial performance toward methicillin‐resistant Staphylococcus aureus (MRSA). Besides, under laser irradiation, Ag‐Bi@SiO₂ NPs at 100 µg mL^?1 can effectively obliterate mature MRSA biofilm and cause a 69.5% decrease in the biomass, showing a better therapeutic effect than Bi@SiO₂ NPs with laser (26.8%) or Ag‐Bi@SiO₂ NPs without laser treatment (30.8%) groups. More importantly, in vivo results confirm that ≈95.4% of bacteria in abscess are killed and the abscess ablation is accelerated using the Ag‐Bi@SiO₂ NPs antibacterial platform. Therefore, Ag‐Bi@SiO₂ NPs with photothermal‐enhanced antibacterial activity are a potential nano‐antibacterial agent for the treatment of skin infections.     

Advanced Biomaterials Accumulation of Citrate‐Coated Magnetic Iron Oxide Nanoparticles by Cultured Brain Astrocytes

Magnetic iron oxide nanoparticles (Fe‐NP) are considered for various applications in the brain. However, little is known so far on the uptake and the metabolism of such nanoparticles in brain cells. Since astrocytes are strategically localized between capillaries and neurons, astrocytes are of particular interest concerning uptake and fate of nanoparticles in the brain. Using astrocyte‐rich primary cultures as model system we have investigated the accumulation of citrate‐coated Fe‐NP by astrocytes. Viable cultured astrocytes accumulate iron from citrate‐coated Fe‐NP in a time‐, concentration‐, and temperature‐dependent manner. The cellular iron content determined after 4 h of incubation increases proportional to the concentration of Fe‐NP, if the particles were applied in concentrations of up to 1000 × 10^?6 M of total iron. The iron accumulation from 500 or 1000 × 10^?6 M iron as Fe‐NP is significantly slowed by lowering the incubation temperature from 37 to 4 °C. Transmission electron microscopy of the cells revealed that most of the cellular Fe‐NP are present in intracellular vesicles. These data demonstrate that astrocytes accumulate efficiently citrate‐coated Fe‐NP, most likely by an endocytotic pathway.     

Biosynthesised AgCl NPs using Bacillus sp. 1/11 and evaluation of their cytotoxic activity and antibacterial and antibiofilm effects on multi‐drug resistant bacteria

Silver nanoparticles (AgNPs) have attracted the attention of researchers due to their properties. Biological synthesis of AgNPs is eco‐friendly and cost‐effective preferred to physical and chemical methods, which utilize environmentally harmful agents and large amounts of energy. Microorganisms have been explored as potential biofactories to synthesize AgNPs. Bacterial NP synthesis is affected by Ag salt concentration, pH, temperature and bacterial species. In this study, Bacillus spp., isolated from soil, were screened for AgNP synthesis at pH 12 with 5 mM Ag nitrate (AgNO₃) final concentration at room temperature. The isolate with fastest color change and the best ultraviolet‐visible spectrum in width and height were chosen as premier one. AgNO₃ and citrate salts were compared in terms of their influence on NP synthesis. Spherical Ag chloride (AgCl) NPs with a size range of 35–40 nm were synthesized in 1.5 mM Ag citrate solution. Fourier transform infrared analysis demonstrated that protein and carbohydrates were capping agents for NPs. In this study, antimicrobial and antitumor properties of the AgNP were investigated. The resulting AgCl NPs had bacteriostatic activity against four standard spp. And multi‐drug resistant strain of Pseudomonas aeruginosa. These NPs are also cytotoxic to cancer cell lines MCF‐7, U87MG and T293.Inspec keywords: silver compounds, nanoparticles, nanomedicine, nanofabrication, particle size, biomedical materials, microorganisms, ultraviolet spectra, visible spectra, Fourier transform infrared spectra, proteins, macromolecules, antibacterial activity, tumours, cancer, cellular biophysics, toxicologyOther keywords: citrate salts, spherical Ag chloride, particle size, Ag citrate solution, Fourier transform infrared analysis, protein, carbohydrates, capping agents, antitumour properties, bacteriostatic activity, Pseudomonas aeruginosa, multidrug resistant strain, cancer cell lines MCF‐7,U87MG, size 35 nm to 40 nm, temperature 293 K to 298 K, AgCl, ultraviolet‐visible spectrum, colour change, room temperature, Ag nitrate final concentration, soil, bacterial species, temperature effect, pH, Ag salt concentration, biofactories, microorganisms, environmentally harmful agents, chemical methods, physical methods, antibacterial properties, electrical properties, mechanical properties, silver nanoparticles, multidrug resistant bacteria, antibiofilm effects, antibacterial effects, cytotoxic activity, Bacillus sp. 1/11, biosynthesised AgCl NPs     

Quantitative assessment of the comparative nanoparticle-uptake efficiency of a range of cell lines

The mechanism(s) of nanoparticle-cell interactions are still not understood. At present there is little knowledge of the relevant length- and timescales for nanoparticle intracellular entry and localization within cells, or the cell-specificity of nanoparticle uptake and localisation. Here, the effect of particle size on the in-vitro intracellular uptake of model fluorescent carboxyl-modified polystyrene nanoparticles is investigated in various cell lines. A range of micro- and nanoparticles of defined sizes (40 nm to 2 μm) are incubated with a series of cell types, including HeLa and A549 epithelial cells, 1321N1 astrocytes, HCMEC D3 endothelial cells, and murine RAW 264.7 macrophages. Techniques such as confocal microscopy and flow cytometry are used to study particle uptake and subcellular localisation, making significant efforts to ensure reproducibility in a semiquantitative approach. The results indicate that internalization of (nano)particles is highly size-dependent for all cell lines studied, and the kinetics of uptake for the same type of nanoparticle varies in the different cell types. Interestingly, even cells not specialized for phagocytosis are able to internalize the larger nanoparticles. Intracellular uptake of all sizes of particles is observed to be highest in RAW 264.7 cells (a specialized phagocytic cell line) and the lowest in the HeLa cells. These results suggest that (nano)particle uptake might not follow commonly defined size limits for uptake processes, and highlight the variability of uptake kinetics for the same material in different cell types. These conclusions have important implications for the assessment of the safety of nanomaterials and for the potential biomedical applications of nanoparticles.     

Multiphoton Microscopy of Nonfluorescent Nanoparticles In Vitro and In Vivo

Nanotechnology holds great promise for a plethora of potential applications. The interaction of engineered nanomaterials with living cells, tissues, and organisms is, however, only partly understood. Microscopic investigations of nano‐bio interactions are mostly performed with a few model nanoparticles (NPs) which are easy to visualize, such as fluorescent quantum dots. Here the possibility to visualize nonfluorescent NPs with multiphoton excitation is investigated. Signals from silver (Ag), titanium dioxide (TiO₂), and silica (SiO₂) NPs in nonbiological environments are characterized to determine signal dependency on excitation wavelength and intensity as well as their signal stability over time. Ag NPs generate plasmon‐induced luminescence decaying over time. TiO₂ NPs induce photoluminescent signals of variable intensities and in addition strong third harmonic generation (THG). Optimal settings for microscopic detection are determined and then applied for visualization of these two particle types in living cells, in murine muscle tissue, and in the murine blood stream. Silica NPs produce a THG signal, but in living cells it cannot be discriminated sufficiently from endogenous cellular structures. It is concluded that multiphoton excitation is a viable option for studies of nano‐bio interactions not only for fluorescent but also for some types of nonfluorescent NPs.