Dual-Mode Mass Spectrometric Imaging for Determination of in Vivo Stability of Nanoparticle Monolayers.

Effective correlation of the in vitro and in vivo stability of nanoparticle-based platforms is a key challenge in their translation into the clinic. Here, we describe a dual imaging method that site-specifically reports the stability of monolayer-functionalized nanoparticles in vivo. This approach uses laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) imaging to monitor the distributions of the nanoparticle core material and laser desorption/ionization mass spectrometry (LDI-MS) imaging to report on the monolayers on the nanoparticles. Quantitative comparison of the images reveals nanoparticle stability at the organ and suborgan level. The stability of particles observed in the spleen was location-dependent and qualitatively similar to in vitro studies. In contrast, in vivo stability of the nanoparticles in the liver differed dramatically from in vitro studies, demonstrating the importance of in vivo assessment of nanoparticle stability.

Nanoparticle-dendrimer hybrid nanocapsules for therapeutic delivery.

BACKGROUND: Nanocapsules can efficiently encapsulate therapeutic cargo for anticancer drug delivery. However, the controlled release of the payload remains a challenge for effective drug delivery. MATERIALS & METHODS: We used dithiocarbamate-functionalized PAMAM dendrimer to cross-link the shell of arginine gold nanoparticles stabilized nanocapsule, and controlled the drug release from the nanocapsule. The ability of cross-linked nanocapsule to deliver hydrophobic paclitaxel to B16F10 cells was demonstrated both in vitro and in vivo. RESULTS: Cross-linked nanocapsule possesses tunable stability and modular permeability, and can deliver paclitaxel with improved anticancer efficiency compared with free drug both in vitro and in vivo. CONCLUSION: Dithiocarbamate chemistry provides a new tool to harness multifactorial colloidal self-assembly for controlled drug delivery for cancer therapy.

Surface Charge Controls the Suborgan Biodistributions of Gold Nanoparticles.

Surface chemistry plays a deciding role in nanoparticle biodistribution, yet very little is known about how surface chemistry influences the suborgan distributions of nanomaterials. Here, using quantitative imaging based on laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS), we demonstrate that surface charge dictates the suborgan distributions of nanoparticles in the kidney, liver, and spleen of mice intravenously injected with functionalized gold nanoparticles. Images of the kidney show that positively charged nanoparticles accumulate extensively in the glomeruli, the initial stage in filtering for the nephron, suggesting that these nanoparticles may be filtered by the kidney at a different rate than the neutral or negatively charged nanoparticles. We find that positively and negatively charged nanoparticles accumulate extensively in the red pulp of the spleen. In contrast, uncharged nanoparticles accumulate in the white pulp and marginal zone of the spleen to a greater extent than the positively or negatively charged nanoparticles. Moreover, these uncharged nanoparticles are also more likely to be found associated with Kupffer cells in the liver. Positively charged nanoparticles accumulate extensively in liver hepatocytes, whereas negatively charged nanoparticles show a broader distribution in the liver. Together these observations suggest that neutral nanoparticles having 2 nm cores may interact with the immune system to a greater extent than charged nanoparticles, highlighting the value of determining the suborgan distributions of nanomaterials for delivery and imaging applications.

Application of Chang's attenuation correction technique for single-photon emission computed tomography partial angle acquisition of Jaszczak phantom.

The acquisition and processing of the Jaszczak phantom is a recommended test by the American College of Radiology for evaluation of gamma camera system performance. To produce the reconstructed phantom image for quality evaluation, attenuation correction is applied. The attenuation of counts originating from the center of the phantom is greater than that originating from the periphery of the phantom causing an artifactual appearance of inhomogeneity in the reconstructed image and complicating phantom evaluation. Chang's mathematical formulation is a common method of attenuation correction applied on most gamma cameras that do not require an external transmission source such as computed tomography, radionuclide sources installed within the gantry of the camera or a flood source. Tomographic acquisition can be obtained in two different acquisition modes for dual-detector gamma camera; one where the two detectors are at 180° configuration and acquire projection images for a full 360°, and the other where the two detectors are positioned at a 90° configuration and acquire projections for only 180°. Though Chang's attenuation correction method has been used for 360° angle acquisition, its applicability for 180° angle acquisition remains a question with one vendor's camera software producing artifacts in the images. This work investigates whether Chang's attenuation correction technique can be applied to both acquisition modes by the development of a Chang's formulation-based algorithm that is applicable to both modes. Assessment of attenuation correction performance by phantom uniformity analysis illustrates improved uniformity with the proposed algorithm (22.6%) compared to the camera software (57.6%).

Regulation of Macrophage Recognition through the Interplay of Nanoparticle Surface Functionality and Protein Corona.

Using a family of cationic gold nanoparticles (NPs) with similar size and charge, we demonstrate that proper surface engineering can control the nature and identity of protein corona in physiological serum conditions. The protein coronas were highly dependent on the hydrophobicity and arrangement of chemical motifs on NP surface. The NPs were uptaken in macrophages in a corona-dependent manner, predominantly through recognition of specific complement proteins in the NP corona. Taken together, this study shows that surface functionality can be used to tune the protein corona formed on NP surface, dictating the interaction of NPs with macrophages.

Quantitative imaging of 2 nm monolayer-protected gold nanoparticle distributions in tissues using laser ablation inductively-coupled plasma mass spectrometry (LA-ICP-MS).

Functionalized gold nanoparticles (AuNPs) have unique properties that make them important biomedical materials. Optimal use of these materials, though, requires an understanding of their fate in vivo. Here we describe the use of laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) to image the biodistributions of AuNPs in tissues from mice intravenously injected with AuNPs. We demonstrate for the first time that the distributions of very small (∼2 nm core) monolayer-protected AuNPs can be imaged in animal tissues at concentrations in the low parts-per-billion range. Moreover, the LA-ICP-MS images reveal that the monolayer coatings on the injected AuNPs influence their distributions, suggesting that the AuNPs remain intact in vivo and their surface chemistry influences how they interact with different organs. We also demonstrate that quantitative images of the AuNPs can be generated when the appropriate tissue homogenates are chosen for matrix matching. Overall, these results demonstrate the utility of LA-ICP-MS for tracking the fate of biomedically-relevant AuNPs in vivo, facilitating the design of improved AuNP-based therapeutics.

Ratiometric Array of Conjugated Polymers-Fluorescent Protein Provides a Robust Mammalian Cell Sensor.

Supramolecular complexes of a family of positively charged conjugated polymers (CPs) and green fluorescent protein (GFP) create a fluorescence resonance energy transfer (FRET)-based ratiometric biosensor array. Selective multivalent interactions of the CPs with mammalian cell surfaces caused differential change in FRET signals, providing a fingerprint signature for each cell type. The resulting fluorescence signatures allowed the identification of 16 different cell types and discrimination between healthy, cancerous, and metastatic cells, with the same genetic background. While the CP-GFP sensor array completely differentiated between the cell types, only partial classification was achieved for the CPs alone, validating the effectiveness of the ratiometric sensor. The utility of the biosensor was further demonstrated in the detection of blinded unknown samples, where 121 of 128 samples were correctly identified. Notably, this selectivity-based sensor stratified diverse cell types in minutes, using only 2000 cells, without requiring specific biomarkers or cell labeling.

Fabrication of Robust Protein Films Using Nanoimprint Lithography.

A nanoimprint-lithography-based fabrication method to generate stable protein films is described. The process is environmentally friendly and generalizable with respect to the protein building blocks. These non-fouling surfaces are readily patternable, incorporate intrinsic protein charge into the film, and able to control cellular adhesion.

A Multichannel Biosensor for Rapid Determination of Cell Surface Glycomic Signatures.

Cell surface glycosylation serves a fundamental role in dictating cell and tissue behavior. Cell surface glycomes differ significantly, presenting viable biomarkers for identifying cell types and their states. Glycoprofiling is a challenging task, however, due to the complexity of the constituent glycans. We report here a rapid and effective sensor for surface-based cell differentiation that uses a three-channel sensor produced by noncovalent conjugation of a functionalized gold nanoparticle (AuNP) and fluorescent proteins. Wild-type and glycomutant mammalian cells were effectively stratified using fluorescence signatures obtained from a single sensor element. Blinded unknowns generated from the tested cell types were identified with high accuracy (44 out of 48 samples), validating the robustness of the multichannel sensor. Notably, this selectivity-based high-throughput sensor differentiated between cells, employing a nondestructive protocol that required only a single well of a microplate for detection.

Binding Studies of Cucurbit[7]uril with Gold Nanoparticles Bearing Different Surface Functionalities.

Host-guest interactions between a synthetic receptor, cucurbit[7]uril (CB[7]), and gold nanoparticles (AuNPs) have been quantified using isothermal titration calorimetry. AuNPs were functionalized with ligands containing tertiary or quaternary benzylamine derivatives, with electron donating or withdrawing groups at the para position of the benzene ring. Analysis of binding interactions reveals that functional groups at the para position have no significant effect on binding constant. However, headgroups bearing a permanent positive charge increased the binding of AuNPs to CB[7] ten-fold compared to monomethyl counterparts.

Acylsulfonamide-Functionalized Zwitterionic Gold Nanoparticles for Enhanced Cellular Uptake at Tumor pH.

A nanoparticle design featuring pH-responsive alkoxyphenyl acylsulfonamide ligands is reported herein. As a result of ligand structure, this nanoparticle is neutral at pH 7.4, becoming positively charged at tumor pH (<6.5). The particle uptake and cytotoxicity increase over this pH range. This pH-controlled uptake and toxicity makes this particle a promising tool for tumor selective therapy.

Fabrication of functional nanofibers through post-nanoparticle functionalization.

A facile method is developed to functionalize nanofiber surfaces with nanoparticles (NPs) through dithiocarbamate chemistry. Gold nanoparticles (AuNPs) and quantum dots (QDs) are immobilized on the nanofiber surface. These surfaces provide scaffolds for further supramolecular functionalization, as demonstrated through the Förster resonance energy transfer (FRET) pairing of QD-decorated fibers and fluorescent proteins.

Probing the Protein-Nanoparticle Interface: The Role of Aromatic Substitution Pattern on Affinity.

A new class of cationic gold nanoparticles has been synthesized bearing benzyl moieties featuring -NO2 and -OMe groups to investigate the regioisomeric control of aromatic nanoparticle-protein recognition. In general, nanoparticles bearing electron withdrawing group demonstrated higher binding affinities towards green fluorescent protein (GFP) compared to electron-donating groups. Significantly, a ~7.5 and ~4.3 fold increase in binding with GFP was observed for -NO2 groups in meta- and para-position respectively, while ortho-substitution showed similar binding compared to the unsubstituted ring. These findings demonstrated that nanoparticle-protein interaction can be controlled by the tuning the spatial orientation and the relative electronic properties of the aromatic substituents. This improved biomolecular recognition provides opportunities for enhanced biosensing and functional protein delivery to the cells.

A multichannel nanosensor for instantaneous readout of cancer drug mechanisms.

Screening methods that use traditional genomic, transcriptional, proteomic and metabonomic signatures to characterize drug mechanisms are known. However, they are time consuming and require specialized equipment. Here, we present a high-throughput multichannel sensor platform that can profile the mechanisms of various chemotherapeutic drugs in minutes. The sensor consists of a gold nanoparticle complexed with three different fluorescent proteins that can sense drug-induced physicochemical changes on cell surfaces. In the presence of cells, fluorescent proteins are rapidly displaced from the gold nanoparticle surface and fluorescence is restored. Fluorescence 'turn on' of the fluorescent proteins depends on the drug-induced cell surface changes, generating patterns that identify specific mechanisms of cell death induced by drugs. The nanosensor is generalizable to different cell types and does not require processing steps before analysis, offering an effective way to expedite research in drug discovery, toxicology and cell-based sensing.

Rapid identification of bacterial biofilms and biofilm wound models using a multichannel nanosensor.

Identification of infectious bacteria responsible for biofilm-associated infections is challenging due to the complex and heterogeneous biofilm matrix. To address this issue and minimize the impact of heterogeneity on biofilm identification, we developed a gold nanoparticle (AuNP)-based multichannel sensor to detect and identify biofilms based on their physicochemical properties. Our results showed that the sensor can discriminate six bacterial biofilms including two composed of uropathogenic bacteria. The capability of the sensor was further demonstrated through discrimination of biofilms in a mixed bacteria/mammalian cell in vitro wound model.

Iterative reconstruction using a Monte Carlo based system transfer matrix for dedicated breast positron emission tomography.

To maximize sensitivity, it is desirable that ring Positron Emission Tomography (PET) systems dedicated for imaging the breast have a small bore. Unfortunately, due to parallax error this causes substantial degradation in spatial resolution for objects near the periphery of the breast. In this work, a framework for computing and incorporating an accurate system matrix into iterative reconstruction is presented in an effort to reduce spatial resolution degradation towards the periphery of the breast. The GATE Monte Carlo Simulation software was utilized to accurately model the system matrix for a breast PET system. A strategy for increasing the count statistics in the system matrix computation and for reducing the system element storage space was used by calculating only a subset of matrix elements and then estimating the rest of the elements by using the geometric symmetry of the cylindrical scanner. To implement this strategy, polar voxel basis functions were used to represent the object, resulting in a block-circulant system matrix. Simulation studies using a breast PET scanner model with ring geometry demonstrated improved contrast at 45% reduced noise level and 1.5 to 3 times resolution performance improvement when compared to MLEM reconstruction using a simple line-integral model. The GATE based system matrix reconstruction technique promises to improve resolution and noise performance and reduce image distortion at FOV periphery compared to line-integral based system matrix reconstruction.

Effect of nano-scale curvature on the intrinsic blood coagulation system.

The intrinsic coagulation activity of silica nanoparticles strongly depends on their surface curvature. Nanoparticles with higher surface curvature do not denature blood coagulation factor XII on its surface, providing a coagulation 'silent' surface, while nanoparticles with lower surface curvature show denaturation and concomitant coagulation.

Functional gold nanoparticles as potent antimicrobial agents against multi-drug-resistant bacteria.

We present the use of functionalized gold nanoparticles (AuNPs) to combat multi-drug-resistant pathogenic bacteria. Tuning of the functional groups on the nanoparticle surface provided gold nanoparticles that were effective against both Gram-negative and Gram-positive uropathogens, including multi-drug-resistant pathogens. These AuNPs exhibited low toxicity to mammalian cells, and bacterial resistance was not observed after 20 generations. A strong structure-activity relationship was observed as a function of AuNP functionality, providing guidance to activity prediction and rational design of effective antimicrobial nanoparticles.

Fabrication of corona-free nanoparticles with tunable hydrophobicity.

A protein corona is formed at the surface of nanoparticles in the presence of biological fluids, masking the surface properties of the particle and complicating the relationship between chemical functionality and biological effects. We present here a series of zwitterionic NPs of variable hydrophobicity that do not adsorb proteins at moderate levels of serum protein and do not form hard coronas at physiological serum concentrations. These particles provide platforms to evaluate nanobiological behavior such as cell uptake and hemolysis dictated directly by chemical motifs at the nanoparticle surface.

Gold nanoparticles for nucleic acid delivery.

Gold nanoparticles provide an attractive and applicable scaffold for delivery of nucleic acids. In this review, we focus on the use of covalent and noncovalent gold nanoparticle conjugates for applications in gene delivery and RNA-interference technologies. We also discuss challenges in nucleic acid delivery, including endosomal entrapment/escape and active delivery/presentation of nucleic acids in the cell.