Monitoring AuNP Dynamics in the Blood of a Single Mouse Using Single Particle Inductively Coupled Plasma Mass Spectrometry with an Ultralow-Volume High-Efficiency Introduction System.

Gold nanoparticles (AuNPs) are increasingly being used as diagnostic and therapeutic agents owing to their excellent properties; however, there is not much data available on their dynamics in vivo on a single particle basis in a single mouse. Here, we developed a method for the direct analysis of nanoparticles in trace blood samples based on single particle inductively coupled plasma-mass spectrometry (spICP-MS). A flexible, highly configurable, and precisely controlled sample introduction system was designed by assembling an ultralow-volume autosampler (flow rate in the range of 5-5000 µL/min) and a customized cyclonic spray chamber (transfer efficiency up to 99%). Upon systematic optimization, the detection limit of the nanoparticle size (LODsize) of AuNPs in ultrapure water was 19 nm, and the detection limit of the nanoparticle number concentration (LODNP) was 8 × 104 particle/L. Using a retro-orbital blood sampling method and subsequent dilution, the system was successfully applied to track the dynamic changes in size and concentration for AuNPs in the blood of a single mouse, and the recovery for the blood sample was 111.74%. Furthermore, the concentration of AuNPs in mouse blood reached a peak in a short period of time and, then, gradually decreased. This study provides a promising technique for analyzing and monitoring the size and concentration of nanoparticles in ultralow-volume blood samples with low concentrations, making it a powerful tool for analyzing and understanding the fate of nanoparticles in vivo.

Surface chemistry governs the sub-organ transfer, clearance and toxicity of functional gold nanoparticles in the liver and kidney.

BACKGROUND: To effectively applied nanomaterials (NMs) in medicine, one of the top priorities is to address a better understanding of the possible sub-organ transfer, clearance routes, and potential toxicity of the NMs in the liver and kidney. RESULTS: Here we explored how the surface chemistry of polyethylene glycol (PEG), chitosan (CS), and polyethylenimine (PEI) capped gold nanoparticles (GNPs) governs their sub-organ biodistribution, transfer, and clearance profiles in the liver and kidney after intravenous injection in mice. The PEG-GNPs maintained dispersion properties in vivo, facilitating passage through the liver sinusoidal endothelium and Disse space, and were captured by hepatocytes and eliminated via the hepatobiliary route. While, the agglomeration/aggregation of CS-GNPs and PEI-GNPs in hepatic Kupffer and endothelial cells led to their long-term accumulation, impeding their elimination. The gene microarray analysis shows that the accumulation of CS-GNPs and PEI-GNPs in the liver induced obvious down-regulation of Cyp4a or Cyp2b related genes, suggesting CS-GNP and PEI-GNP treatment impacted metabolic processes, while the PEI-GNP treatment is related with immune responses. CONCLUSIONS: This study demonstrates that manipulation of nanoparticle surface chemistry can help NPs selectively access distinct cell types and elimination pathways, which help to clinical potential of non-biodegradable NPs.

Phase I Clinical Trial Results of Auranofin, a Novel Antiparasitic Agent.

Under an NIH priority to identify new drugs to treat class B parasitic agents, we performed high-throughput screens, which identified the activity of auranofin (Ridaura) against Entamoeba histolytica and Giardia intestinalis, major causes of water- and foodborne outbreaks. Auranofin, an orally administered, gold (Au)-containing compound that was approved by the FDA in 1985 for treatment of rheumatoid arthritis, was effective in vitro and in vivo against E. histolytica and both metronidazole-sensitive and -resistant strains of Giardia We now report the results of an NIH-sponsored phase I trial to characterize the pharmacokinetics (PK) and safety of auranofin in healthy volunteers using modern techniques to measure gold levels. Subjects received orally 6 mg (p.o.) of auranofin daily, the recommended dose for rheumatoid arthritis, for 7 days and were followed for 126 days. Treatment-associated adverse events were reported by 47% of the subjects, but all were mild and resolved without treatment. The mean gold maximum concentration in plasma (Cmax) at day 7 was 0.312 µg/ml and the half-life (t1/2) 35 days, so steady-state blood levels would not be reached in short-term therapy. The highest concentration of gold, 13 µM (auranofin equivalent), or more than 25× the 50% inhibitory concentration (IC50) for E. histolytica and 4× that for Giardia, was in feces at 7 days. Modeling of higher doses (9 and 21 mg/day) was performed for systemic parasitic infections, and plasma gold levels of 0.4 to 1.0 µg/ml were reached after 14 days of treatment at 21 mg/day. This phase I trial supports the idea of the safety of auranofin and provides important PK data to support its potential use as a broad-spectrum antiparasitic drug. (This study has been registered at ClinicalTrials.gov under identifier NCT02089048.).

Gold nanoparticle interactions in human blood: a model evaluation.

In this study, we investigated gold nanoparticle (AuNP) interactions in blood using thromboelastography as a rapid screening tool to monitor their influence on blood coagulation. 1.2 nM colloidal AuNPs ranging from 12 to 85 nm have no effect in the blood, however, 5 nM AuNPs demonstrate pro-thrombogenic concentration dependent effects with a reduction in clot formation. Size effects exhibit a non-linear trend with 45 and 85 nm particles resulting in a faster pro-thrombotic response. Clot strength decreased with AuNP size with the greatest reduction with 28 nm particles. We assessed AuNP interactions in the blood focusing on their biological activity. AuNP-RGD possessed pro-coagulant activities, while PEG-thiol, human fibrinogen and clopidogrel prevented blood clot formation and influenced platelet activity, and were more efficient when bound to nanocarriers than unbound ligands. Such tests could fill the knowledge gaps in thrombogenicity of NPs between in vitro test methods and predict in vivo haemocompatibility.

Single Particle Plasmon Sensors as Label-Free Technique To Monitor MinDE Protein Wave Propagation on Membranes.

We use individual gold nanorods as pointlike detectors for the intrinsic dynamics of an oscillating biological system. We chose the pattern forming MinDE protein system from Escherichia coli (E. coli), a prominent example for self-organized chemical oscillations of membrane-associated proteins that are involved in the bacterial cell division process. Similar to surface plasmon resonance (SPR), the gold nanorods report changes in their protein surface coverage without the need for fluorescence labeling, a technique we refer to as NanoSPR. Comparing the dynamics for fluorescence labeled and unlabeled proteins, we find a reduction of the oscillation period by about 20%. The absence of photobleaching allows us to investigate Min proteins attaching and detaching from lipid coated gold nanorods with an unprecedented bandwidth of 100 ms time resolution and 1 h observation time. The long observation reveals small changes of the oscillation period over time. Averaging many cycles yields the precise wave profile that exhibits the four phases suggested in previous reports. Unexpected from previous fluorescence-based studies, we found an immobile static protein layer not dissociating during the oscillation cycle. Hence, NanoSPR is an attractive label-free real-time technique for the local investigation of molecular dynamics with high observation bandwidth. It gives access to systems, which cannot be fluorescently labeled, and resolves local dynamics that would average out over the sensor area used in conventional SPR.

Gold nanoparticles as a substrate in bio-analytical near-infrared surface-enhanced Raman spectroscopy.

As biospectroscopy techniques continue to be developed for screening or diagnosis within a point-of-care setting, an important development for this field will be high-throughput optimization. For many of these techniques, it is therefore necessary to adapt and develop parameters to generate a robust yet simple approach delivering high-quality spectra from biological samples. Specifically, this is important for surface-enhanced Raman spectroscopy (SERS) wherein there are multiple variables that can be optimised to achieve an enhancement of the Raman signal from a sample. One hypothesis is that "large" diameter (>100 nm) gold nanoparticles provide a greater enhancement at near-infrared (NIR) and infrared (IR) wavelengths than those <100 nm in diameter. Herein, we examine this notion using examples in which SERS spectra were acquired from MCF-7 breast cancer cells incubated with 150 nm gold nanoparticles. It was found that 150 nm gold nanoparticles are an excellent material for NIR/IR SERS. Larger gold nanoparticles may better satisfy the theoretical restraints for SERS enhancement at NIR/IR wavelengths compared to smaller nanoparticles. Also, larger nanoparticles or their aggregates are more readily observed via optical microscopy (and especially electron microscopy) compared to smaller ones. This allows rapid and straightforward identification of target areas containing a high concentration of nanoparticles and facilitating SERS spectral acquisition. To some extent, these observations appear to extend to biofluids such as blood plasma or (especially) serum; SERS spectra of such biological samples often exhibit a low signal-to-noise ratio in the absence of nanoparticles. With protein-rich biofluids such as serum, a dramatic SERS effect can be observed; although this might facilitate improved spectral biomarker identification in the future, it may not always improve classification between control vs. cancer. Thus, use of "large" gold nanoparticles are a good starting point in order to derive informative NIR/IR SERS analysis of biological samples.

Biologically inspired stealth peptide-capped gold nanoparticles.

Introduction into the human body makes most nanoparticle systems susceptible to aggregation via nonspecific protein binding. Here, we developed a peptide-capped gold nanoparticle platform that withstands aggregation in undiluted human serum at 37 °C for 24 h. This biocompatible and natural system is based on mimicking human proteins which are enriched in negatively charged glutamic acid and positively charged lysine residues on their surface. The multifunctional EKEKEKE-PPPPC-Am peptide sequence consists of a stealth glutamic acid/lysine portion combined with a surface anchoring linker containing four prolines and a cysteine. Particle stability was measured via optical spectroscopy and dynamic light scattering in single protein, high salt, and undiluted human serum solutions. In vitro cell experiments demonstrate EKEKEKE-PPPPC-Am capped gold nanoparticles effectively minimize nonspecific cell uptake by nonphagocytic bovine aortic endothelial cells and phagocytic murine macrophage RAW 264.7 cells. Cytotoxicity studies show that peptide-capped gold nanoparticles do not affect cell viability. Finally, the peptide EKEKEKE-PPPPC-Am was extended with cyclic RGD to demonstrate specific cell targeting and stealth without using poly(ethylene glycol). Adding the functional peptide via peptide sequence extension avoids complex conjugation chemistries that are used for connection to synthetic materials. Inductively coupled plasma mass spectroscopy results indicate high aortic bovine endothelial cell uptake of c[RGDfE(SGG-KEKEKE-PPPPC-Am)] capped gold nanoparticles and low uptake of the control scrambled sequence c[RDGfE(SGG-KEKEKE-PPPPC-Am)] capped gold nanoparticles.

Functional gold nanoparticles coupled with microporous membranes: a flow controlled assay for colorimetric visualization of proteins.

We report a rapid and simple assay for colorimetric visualization of thrombin at nanomolar levels using functional gold nanoparticles (FAuNPs) coupled with microporous membranes. We used a 29-mer thiolated-thrombin-binding-aptamer (TBA29) to prepare TBA29 functionalized AuNPs (TBA29-AuNPs) for the selective detection of human thrombin. The sensing mechanism is based on the principle of TBA29-AuNPs flowing down through the nitrocellulose membrane (NCM) pores at different flow rates after binding to thrombin. Compared with free TBA29-AuNPs, when thrombin-TBA29-AuNPs were dropped on the NCM, the particles flowed down more easily through the NCM pores along with the buffer solution due to the increase in the gravity of particles. Therefore, color intensities of TBA29-AuNPs on the NCM depended on the concentration of thrombin; the color intensity was lighter when the concentration of thrombin was higher. Thrombin can be detected at the nanomolar level with the naked eye using this colorimetric probe. A protein G modified AuNP based probe (PG-AuNPs/NCM) was employed to detect human immunoglobulin G (hIgG) in plasma samples to demonstrate the practicality of our sensing system. Also, fibrinogen modified Au NPs were analyzed to demonstrate that this concept of detection could be extended to other proteins or systems, by functionalizing with suitable molecules.

Phase I dose escalation study of the PKCι inhibitor aurothiomalate for advanced non-small-cell lung cancer, ovarian cancer, and pancreatic cancer.

Protein kinase C iota (PKCι) is overexpressed in non-small-cell lung cancer, ovarian, and pancreatic cancers, where it plays a critical role in oncogenesis. The gold compound aurothiomalate (ATM) has been shown to inhibit PKCι signaling and exerts potent antitumor activity in preclinical models. We sought to determine the maximum tolerated dose (MTD) of ATM. We conducted a phase I dose escalation trial of ATM in patients with non-small-cell lung cancer, ovarian or pancreatic cancer. Patients received ATM intramuscularly weekly for three cycles (cycle duration 4 weeks) at 25, 50, or 75 mg in a 3+3 design. The dose was not escalated for individual patients. Blood samples were analyzed for elemental gold levels. Patients were evaluated every 4 weeks for toxicity and every 8 weeks for response. Fifteen patients were enrolled in this study. Six patients were treated at 25 mg, seven at 50 mg, and two at 75 mg. There was one dose-limiting toxicity at 25 mg (hypokalemia), one at 50 mg (urinary tract infection), and none at 75 mg. There were three grade 3 hematologic toxicities. The recommended MTD of ATM is 50 mg. Patients received treatment for a median of two cycles (range 1-3). There appeared to be a dose-related accumulation of steady-state plasma concentrations of gold consistent with linear pharmacokinetics. In summary, this phase I study was successful in identifying ATM 50 mg intramuscularly weekly as the MTD. Future clinical investigations targeting PKCι are currently in progress.

In vivo toxicity, biodistribution, and clearance of glutathione-coated gold nanoparticles.

Gold nanoparticles are emerging as promising materials from which to construct nanoscale therapeutics and therapeutic delivery systems. However, animal studies have shown that gold nanoparticles modified with certain thiol monolayers such as tiopronin can cause renal complications and morbidity. Although these effects may be eliminated by coadsorbing small amounts of polyethylene glycol (PEG) onto the nanoparticle surface, PEG can also lower cellular internalization efficiency and binding interactions with protein disease targets, significantly reducing the potential for using gold nanoparticles as therapeutics. Using ICP-MS analysis of blood, urine, and several organs, we show in this article that glutathione-coated gold nanoparticles (1.2 nm ± 0.9 nm) cause no morbidity at any concentration up to and including 60 µM and target primary organs although providing gradual dissipation and clearance over time. This study suggests that glutathione may be an attractive alternative to PEG in the design of gold nanoparticle therapeutics. FROM THE CLINICAL EDITOR: This study describes the utility and toxicity of glutathione coated gold nanoparticles in comparison to PEGylated counterparts that are commonly used to increase "Stealth" properties and lower cytotoxicity. Too much PEG on the NPs can lead to lower cellular internalization efficiency and less efficient binding interactions with protein disease targets, significantly reducing the potential for using gold nanoparticles as therapeutics.

Mixed charged zwitterionic self-assembled monolayers as a facile way to stabilize large gold nanoparticles.

Here we report a facile way of stabilizing large gold nanoparticles (AuNPs) by mixed charged zwitterionic self-assembled monolayers (SAMs). The citrate-capped AuNPs with diameters ranging from 16 nm to even ∼100 nm are well stabilized via a simple place exchange reaction with a 1:1 molar ratio mixture of negatively charged sodium 10-mercaptodecanesulfonic acid (HS-C10-S) and positively charged (10-mercaptodecyl)-trimethyl-ammonium bromide (HS-C10-N4). The 16 nm AuNPs protected by mixed charged zwitterionic SAMs not only show much better stability than the single negatively or positively charged AuNPs, but also exhibit exciting stability as well as those modified by monohydroxy (1-mercaptoundec-11-yl) tetraethylene glycol (HS-C11-EG4). Importantly, 16 nm AuNPs protected by mixed SAMs exhibit good stability in cell culture medium with 10% FBS and strong protein resistance, especially with excellent resistance against plasma adsorption. Moreover, the mixed charged zwitterionic SAMs are also able to well-stabilize larger AuNPs with a diameter of 50 nm, and to help remarkably improve their stability in saline solution compared with HS-C11-EG4 protected ones. When it comes to AuNPs with a diameter of 100 nm, the mixed charged zwitterionic SAM protected nanoparticles retain a smaller hydrodynamic diameter and even better long-term stability than those modified by mercaptopolyethylene glycol (M(w) = 2000, HS-PEG2000). The above results demonstrated that the mixed charged zwitterionic SAMs are able to have a similar effect on stabilizing the large gold nanoparticles just like the single-component zwitterionic SAMs. Concerning its ease of preparation, versatility, and excellent properties, the strategy based on the mixed charged zwitterionic SAM protection might provide a promising method to surface tailoring of nanoparticles for biomedical application.

Se-modified gold nanorods for enhancing the efficiency of photothermal therapy: avoiding the off-target problem induced by biothiols.

Tumor-targeting gold nanorods (AuNRs) assembled through Au-S bonds have been widely used for photothermal therapy (PTT) via intravenous injection. However, with extended in vivo circulation times, biothiols can replace some S-modified targeting ligands on the surface of the AuNRs, which lowers their targeting efficacy towards cancer cells, resulting in a non-ideal PTT effect. To address this problem, herein, we utilized Se-modified AuNRs to establish a dual functional nanoprobe (Casp-RGD-Se-AuNRs) for improving the therapeutic effect and real-time monitoring of Caspase-9 levels to indicate the degree of cell apoptosis. The experiments demonstrated that the Casp-RGD-Se-AuNRs are better at avoiding interference from biothiols than the S-modified nanoprobe (Casp-RGD-S-AuNRs) for extended blood-circulation times after intravenous injection, significantly improving the PTT efficacy via more effectively targeting cancer cells. Simultaneously, the change of Caspase-9 levels visually shows the degree of apoptosis. Moreover, an in vivo study showed that, compared with the S-modified nanoprobe, the Se-modified nanoprobe exhibits a higher delivery efficiency to the tumor region after intravenous injection (accumulation in the tumor increased by 87%) and a better anticancer efficacy under NIR light irradiation (the tumor inhibition rate increased 6-fold). This work provides a valuable strategy to overcome the off-target problem, and new ideas for avoiding interference by biomolecules during blood circulation.

Three-wavelength murine photoplethysmography for estimation of vascular gold nanorod concentration.

Nanoparticle-assisted photo-thermal (NAPT) ablation has become a new and attractive modality for the treatment of cancerous tumors. This therapy exploits the passive accumulation of intravenously delivered optically resonant metal nanoparticles into tumors, however, the circulating bioavailability of these particles is often unknown. We present a non-invasive optical device capable of monitoring the circulation of optically resonant gold nanorods. The device, referred to as a pulse photometer, uses the technique of multi-wavelength photoplethysmography. We simultaneously report the circulation of gold nanorods and oximetry for six hours post-injection in mice with no anesthesia and remove the probe when not collecting data. The instrument shows good agreement (R(2) = 0.903, n = 30) with ex vivo spectrophotometric analysis of blood samples. The real-time feedback provided has a strong potential for reducing variability and thus improving the efficacy of similar clinical therapies.

Unexpected toxicity of monolayer protected gold clusters eliminated by PEG-thiol place exchange reactions.

Monolayer protected clusters (MPCs) are small, metal nanoparticles capped with thiolate ligands that have been widely studied for their size-dependent properties and for their ability to be functionalized for biological applications. Common water-soluble MPCs, functionalized by N-(2-Mercaptopropionyl)-glycine (tiopronin) or glutathione, have been used previously to interface with biological systems. These MPCs are ideal for biological applications not only due to their water-solubility but also their small size (<5 nm). These characteristics are expected to enable easy biodistribution and clearance. In this article, we show an unexpected toxicity is associated with the tiopronin monolayer protected cluster (TMPC), making it incompatible for potential in vivo applications. This toxicity is linked to significant histological damage to the renal tubules, causing mortality at concentrations above 20 µM. We further show how the incorporation of poly ethylene glycol (PEG) by a simple place-exchange reaction eliminates this toxicity. We analyzed gold content within blood and urine and found an increased lifetime of the particle within the bloodstream due to the creation of the mixed monolayer. Also shown was the elimination of kidney damage with the use of the mixed-monolayer particle via Multistix analysis, MALDI-TOF MS analysis, and histological examination. Final immunological analysis showed no effect on white blood cell (WBC) count for the unmodified particle and a surprising increase in WBC count with the injection of mixed monolayer particles at concentrations higher than 30 µM, suggesting that there may be an immune response to these mixed monolayer nanoparticles at high concentrations; therefore, special attention should be focused on selecting the best capping ligands for use in vivo. These findings make the mixed monolayer an excellent candidate for further biological applications using water-soluble nanoparticles.

Contact allergy to gold as a model for clinical-experimental research.

The high frequency of contact allergy to gold in patients with dermatitis was established after exhaustive skin testing, determining the right test agent, the best concentration, and repeated test readings. Metallic gold in contact with skin is slowly ionized, permitting absorption and haptenisation. Contact allergy to gold is statistically correlated to the presence of dental gold. But in many case reports it has also been attributed to wearing gold jewellery, albeit not statistically demonstrated. Epicutaneous testing with gold salts increases the blood gold level, and by intramuscular injection systemic contact dermatitis is provoked in an allergic individual. In coronary heart disease, gold-coated intravascular stents have been shown to be correlated to contact allergy and even to an increased risk of restenosis. Gold is far from inert.

ICP-MS trace element analysis in serum and whole blood.

Trace elements and minerals are compounds that are essential for the support of a variety of biological functions and play an important role in the formation of and the defense against oxidative stress. Here we describe a technique, allowing sequential detection of the trace elements (K, Zn, Se, Cu, Mn, Fe, Mg) in serum and whole blood by an ICP-MS method using single work-up, which is a simple, quick and robust method for the sequential measurement and quantification of the trace elements Sodium (Na), Potassium (K), Calcium (Ca), Zinc (Zn), Selenium (Se), Copper (Cu), Iron (Fe), Manganese (Mn) and Magnesium (Mg) in whole blood as well as Copper (Cu), Selenium (Se), Zinc (Zn), Iron (Fe), Magnesium (Mg), Manganese (Mn), Chromium (Cr), Nickel (Ni), Gold (Au) and Lithium (Li) in human serum. For analysis, only 100 µl of serum or whole blood is sufficient, which make this method suitable for detecting trace element deficiency or excess in newborns and infants. All samples were processed and analyzed by ICP-MS (Agilent Technologies). The accuracy, precision, linearity and the limit of quantification (LOQ), Limit of Blank (LOB) and the limit of detection (LOD) of the method were assessed. Recovery rates were between 80-130% for most of the analyzed elements; repeatabilities (Cv %) calculated were below 15% for most of the measured elements. The validity of the proposed methodology was assessed by analyzing a certified human serum and whole blood material with known concentrations for all elements; the method described is ready for routine use in biomonitoring studies.

SEC hyphenated to a multielement-specific detector unravels the degradation pathway of a bimetallic anticancer complex in human plasma.

The bimetallic metal complex Titanocref exhibits relevant anticancer activity, but it is unknown if it is stable to reach target tissues intact. To gain insight, a pharmacologically relevant dose was added to human blood plasma and the mixture was incubated at 37 °C. The obtained mixture was analyzed 5 and 60 min later by size-exclusion chromatography hyphenated to an inductively coupled plasma atomic emission spectrometer (SEC-ICP-AES). We simultaneously detected several titanium (Ti), gold (Au) and sulfur (S)-peaks, which corresponded to a Ti degradation product that eluted partially, and a Au degradation product that eluted entirely bound to plasma proteins (both time points). Although ~70% of the intact Titanocref was retained on the column after 60 min, our results allowed us to establish - for the first time - its likely degradation pathway in human plasma at near physiological conditions. These results suggest that ~70% of Titanocref remain in plasma after 60 min, which supports results from a recent in vivo study in which mice were treated with Titanocref and revealed Ti:Au molar ratios in tumors and organs close to 1:1. Thus, our stability studies suggest that the intact drug is able to reach target tissue. Overall, our results exemplify that SEC-ICP-AES enables the execution of intermediate in vitro studies with human plasma in the context of advancing bimetallic metal-based drugs to more costly clinical studies.

A Systematic comparison of in vitro cell uptake and in vivo biodistribution for three classes of gold nanoparticles with saturated PEG coatings.

A great deal of attention has been focused on nanoparticles for cancer therapy, with the promise of tumor-selective delivery. However, despite intense work in the field over many years, the biggest obstacle to this vision remains extremely low delivery efficiency of nanoparticles into tumors. Due to the cost, time, and impact on the animals for in vivo studies, the nanoparticle field predominantly uses cellular uptake assays as a proxy to predict in vivo outcomes. Extensive research has focused on decreasing macrophage uptake in vitro as a proxy to delay nanoparticle accumulation in the mononuclear phagocytic system (MPS), mainly the liver and spleen, and thereby increase tumor accumulation. We have recently reported novel synthetic methods employing small molecule crosslinkers for the controlled assembly of small nanoparticles into larger aggregates and found that these nanoaggregates had remarkably high surface coverage and low cell uptake, even in macrophages. We further found that this extremely low cellular uptake could be recapitulated on solid gold nanoparticles by densely coating their surface with small molecules. Here we report our studies on the biodistribution and clearance of these materials in comparison to more conventional PEGylated gold nanoparticles. It was expected that the remarkably low macrophage uptake in vitro would translate to extended blood circulation time in vivo, but instead we found no correlation between either surface coverage or in vitro macrophage cell uptake and in vivo blood circulation. Gold nanoaggregates accumulate more rapidly and to a higher level in the liver compared to control gold nanoparticles. The lack of correlation between in vitro macrophage uptake and in vivo blood circulation suggests that the field must find other in vitro assays to use as a primary proxy for in vivo outcomes or use direct in vivo experimentation as a primary assay.

Does gold concentration in the blood influence the result of patch testing to gold?

BACKGROUND: We have recently found a correlation between contact allergy to gold sodium thiosulphate (GSTS) and gold concentration in the blood (B-Au) in a stented population: the higher the B-Au, the stronger the patch-test reaction. OBJECTIVES: To further investigate the correlation between B-Au and patch-test reactivity to gold. METHODS: In this provocation control cross-over trial of 24 patients with dermatitis with a known contact allergy to gold, the patients were randomized into two groups where one was topically provoked to gold (15 mg GSTS) and one to the control. All patients were simultaneously patch tested with GSTS in 10 aqueous dilutions (1.1 mg GSTS). Patch-test readings were performed and blood was drawn. After 6 weeks, the experiment was repeated and the group that had previously been provoked with gold was now provoked with the control and vice versa. RESULTS: B-Au was higher after gold provocation whereas no treatment effect was discerned for minimal eliciting concentration (MEC) or summarized test score (STS). Instead, significant differences in period effect were observed implying higher B-Au and STS and lower MEC on test occasion II. The most likely explanation is the increased B-Au and /or booster effect from test occasion I. There was a correlation between B-Au and MEC: the higher the B-Au, the lower the MEC. CONCLUSIONS: The correlation between B-Au and MEC indicates that the B-Au is of importance for the skin reactivity to gold.

Proteomic analysis of the bio-corona formed on the surface of (Au, Ag, Pt)-nanoparticles in human serum.

Adsorption of biomolecules onto nanoparticles surface in biological samples led to the formation of a bio-corona, it could modified the "identity" of nanoparticles, contributing to the determination of their toxicity and biocompatibility. Gel electrophoresis in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed to qualitatively analyze and identify the human serum proteins adsorbed on the surface of three different nanomaterials stabilized with citrate: 10.02 ± 0.91 nm gold nanoparticles (AuNPs), 9.73 ± 1.70 nm silver nanoparticles (AgNPs) and, 2.40 ± 0.30 nm platinum nanoparticles (PtNPs). An exhaustive analysis and classification of all identified proteins related with their function were also developed.