Supervised Learning and Mass Spectrometry Predicts the in Vivo Fate of Nanomaterials.

The surface of nanoparticles changes immediately after intravenous injection because blood proteins adsorb on the surface. How this interface changes during circulation and its impact on nanoparticle distribution within the body is not understood. Here, we developed a workflow to show that the evolution of proteins on nanoparticle surfaces predicts the biological fate of nanoparticles in vivo. This workflow involves extracting nanoparticles at multiple time points from circulation, isolating the proteins off the surface and performing proteomic mass spectrometry. The mass spectrometry protein library served as inputs, while blood clearance and organ accumulation were used as outputs to train a supervised deep neural network that predicts nanoparticle biological fate. In a double-blinded study, we tested the network by predicting nanoparticle spleen and liver accumulation with upward of 94% accuracy. Our neural network discovered that the mechanism of liver and spleen uptake is due to patterns of a multitude of nanoparticle surface adsorbed proteins. There are too many combinations to change these proteins manually using chemical or biological inhibitors to alter clearance. Therefore, we developed a technique that uses the host to act as a bioreactor to prepare nanoparticles with predictable clearance patterns that reduce liver and spleen uptake by 50% and 70%, respectively. These techniques provide opportunities to both predict nanoparticle behavior and also to engineer surface chemistries that are specifically designed by the body.

Synthesis of Patient-Specific Nanomaterials.

Nanoparticles are engineered from materials such as metals, polymers, and different carbon allotropes that do not exist within the body. Exposure to these exogenous compounds raises concerns surrounding toxicity, inflammation, and immune activation. These responses could potentially be mitigated by synthesizing nanoparticles directly from molecules derived from the host. However, efforts to assemble patient-derived macromolecules into structures with the same degree of size and shape tunability as their exogenous counterparts remains a significant challenge. Here we solve this problem by creating a new class of size- and shape-tunable personalized protein nanoparticles (PNP) made entirely from patient-derived proteins. PNPs are built into different sizes and shapes with the same degree of tunability as gold nanoparticles. They are biodegradable and do not activate innate or adaptive immunity following single and repeated administrations in vivo. PNPs can be further modified with specific protein cargos that remain catalytically active even after intracellular delivery in vivo. Finally, we demonstrate that PNPs created from different human patients have unique molecular fingerprints encoded directly into the structure of the nanoparticle. This new class of personalized nanomaterial has the potential to revolutionize how we treat patients and can become an integral component in the diagnostic and therapeutic toolbox.

The Role of Nanoparticle Design in Determining Analytical Performance of Lateral Flow Immunoassays.

Rapid, simple, and cost-effective diagnostics are needed to improve healthcare at the point of care (POC). However, the most widely used POC diagnostic, the lateral flow immunoassay (LFA), is ∼1000-times less sensitive and has a smaller analytical range than laboratory tests, requiring a confirmatory test to establish truly negative results. Here, a rational and systematic strategy is used to design the LFA contrast label (i.e., gold nanoparticles) to improve the analytical sensitivity, analytical detection range, and antigen quantification of LFAs. Specifically, we discovered that the size (30, 60, or 100 nm) of the gold nanoparticles is a main contributor to the LFA analytical performance through both the degree of receptor interaction and the ultimate visual or thermal contrast signals. Using the optimal LFA design, we demonstrated the ability to improve the analytical sensitivity by 256-fold and expand the analytical detection range from 3 log10 to 6 log10 for diagnosing patients with inflammatory conditions by measuring C-reactive protein. This work demonstrates that, with appropriate design of the contrast label, a simple and commonly used diagnostic technology can compete with more expensive state-of-the-art laboratory tests.

Peptide-MHC-based nanomedicines for autoimmunity function as T-cell receptor microclustering devices.

We have shown that nanoparticles (NPs) can be used as ligand-multimerization platforms to activate specific cellular receptors in vivo. Nanoparticles coated with autoimmune disease-relevant peptide-major histocompatibility complexes (pMHC) blunted autoimmune responses by triggering the differentiation and expansion of antigen-specific regulatory T cells in vivo. Here, we define the engineering principles impacting biological activity, detail a synthesis process yielding safe and stable compounds, and visualize how these nanomedicines interact with cognate T cells. We find that the triggering properties of pMHC-NPs are a function of pMHC intermolecular distance and involve the sustained assembly of large antigen receptor microclusters on murine and human cognate T cells. These compounds show no off-target toxicity in zebrafish embryos, do not cause haematological, biochemical or histological abnormalities, and are rapidly captured by phagocytes or processed by the hepatobiliary system. This work lays the groundwork for the design of ligand-based NP formulations to re-program in vivo cellular responses using nanotechnology.

Yeast Populations Evolve to Resist CdSe Quantum Dot Toxicity.

Engineered nanomaterials are used globally in biomedical, electronic, and optical devices, and are often discarded into the environment. Cell culture experiments have shown that many inorganic nanoparticles are toxic to eukaryotic cells. Here, we show that populations of eukaryotic cells can evolve to survive chronic exposure to toxic CdSe semiconductor quantum dots (QDs). We grew yeast Saccharomyces cerevisiae for 24 days in liquid medium containing QDs prepared daily at half the minimum inhibitory concentration (MIC50) of the progenitor yeast cells. After 24 days, the cells grew normally under constant exposure to QDs. We concluded that these cells evolved to resist QD toxicity. Surprisingly, when we removed QDs from the growth medium, some of the evolved cells grew poorly, i.e., they grew better in the presence of QDs. Finally, genetic analysis confirmed that the ubiquitin ligase gene bul1 was mutated in the evolved cells, which suggests that this gene may be implicated in increased CdSe QD tolerance. This study shows that chronic exposure to QDs can exert selective pressure causing irreversible genetic changes leading to adaptation.

Engineering the Structure and Properties of DNA-Nanoparticle Superstructures Using Polyvalent Counterions.

DNA assembly of nanoparticles is a powerful approach to control their properties and prototype new materials. However, the structure and properties of DNA-assembled nanoparticles are labile and sensitive to interactions with counterions, which vary with processing and application environment. Here we show that substituting polyamines in place of elemental counterions significantly enhanced the structural rigidity and plasmonic properties of DNA-assembled metal nanoparticles. These effects arose from the ability of polyamines to condense DNA and cross-link DNA-coated nanoparticles. We further used polyamine wrapped DNA nanostructures as structural templates to seed the growth of polymer multilayers via layer-by-layer assembly, and controlled the degree of DNA condensation, plasmon coupling efficiency, and material responsiveness to environmental stimuli by varying polyelectrolyte composition. These results highlight counterion engineering as a versatile strategy to tailor the properties of DNA-nanoparticle assemblies for various applications, and should be applicable to other classes of DNA nanostructures.

Protein corona fingerprinting predicts the cellular interaction of gold and silver nanoparticles.

Using quantitative models to predict the biological interactions of nanoparticles will accelerate the translation of nanotechnology. Here, we characterized the serum protein corona 'fingerprint' formed around a library of 105 surface-modified gold nanoparticles. Applying a bioinformatics-inspired approach, we developed a multivariate model that uses the protein corona fingerprint to predict cell association 50% more accurately than a model that uses parameters describing nanoparticle size, aggregation state, and surface charge. Our model implicates a set of hyaluronan-binding proteins as mediators of nanoparticle-cell interactions. This study establishes a framework for developing a comprehensive database of protein corona fingerprints and biological responses for multiple nanoparticle types. Such a database can be used to develop quantitative relationships that predict the biological responses to nanoparticles and will aid in uncovering the fundamental mechanisms of nano-bio interactions.

Principles of conjugating quantum dots to proteins via carbodiimide chemistry.

The covalent coupling of nanomaterials to bio-recognition molecules is a critical intermediate step in using nanomaterials for biology and medicine. Here we investigate the carbodiimide-mediated conjugation of fluorescent quantum dots to different proteins (e.g., immunoglobulin G, bovine serum albumin, and horseradish peroxidase). To enable these studies, we developed a simple method to isolate quantum dot bioconjugates from unconjugated quantum dots. The results show that the reactant concentrations and protein type will impact the overall number of proteins conjugated onto the surfaces of the quantum dots, homogeneity of the protein-quantum dot conjugate population, quantum efficiency, binding avidity, and enzymatic kinetics. We propose general principles that should be followed for the successful coupling of proteins to quantum dots.

Self-assembly of [60]fullerene-thiol derivatives on mercury surfaces.

Herein we report the first self-assembly of fullerene-thiol conjugates (1 and 2) on thin mercury films (TMF) deposited on a glassy carbon electrode (GCE). The fullerene-containing SAMs were investigated by cyclic voltammetry and water contact angle measurements. Two reversible, surface-confined redox couples were obtained for the fullerene-containing SAMs on TMF/GCE in CH(2)Cl(2) solution. The surface coverage of both fullerene derivatives 1 and 2 on TMF/GCE was measured to be in the range of (1.7-1.8) x 10(-10) mol cm(-2). Both SAMs of 1 and 2 partially blocked the electron transfer across the electrode in aqueous solution. The contact angle measurements performed on TMFs clearly showed an enhancement of the surface hydrophobicity upon formation of the fullerene-containing monolayer.

Photophysical and electrochemical properties of meso, meso-linked oligoporphyrin rods with appended fullerene terminals.

The electrochemical and photophysical properties of molecular architectures consisting of oligomeric meso,meso-linked oligoporphyrin rods linked at both extremities to methanofullerene moieties are presented in comparison to those of model systems. Cyclic voltammetry data evidence the presence of a strong intramolecular electronic coupling along the porphyrin oligomers that varies slightly with their length. This interaction affects the redox potentials of both fullerene and porphyrin moieties. The electronic coupling between the two chromophores is confirmed by comparing the redox potentials of porphyrin arrays before and after attachment of the carbon sphere. Electronic absorption, fluorescence, and phosphorescence spectra of the porphyrin oligomers in toluene are reported, which provide the energy of the lowest singlet and triplet electronic excited states. In the fullerene-porphyrin conjugates, ground-state charge-transfer (CT) interactions are evidenced by low-energy absorption features above 750 nm. These systems also exhibit near-infrared (NIR) CT luminescence in toluene with lifetimes shorter than 1000 ps. On increasing the solvent polarity (from toluene to Et2O and THF), CT emissions become progressively weaker, red-shifted, and shorter lived, which reflects the energy-gap law and Marcus inverted region effects. Luminescence is not detected in benzonitrile. Picosecond transient absorption spectroscopy of the porphyrin-fullerene conjugates allows detection of the porphyrin cation as a clear fingerprint for electron transfer. The rate of charge recombination is in agreement with CT luminescence lifetimes, which confirms the occurrence of NIR radiative back-electron transfer.

A new fullerene complexation ligand: N-pyridylfulleropyrrolidine.

The subject of this paper is a new fullerene building block design with the potential for defined geometry and good electronic communication. The synthesis and characterization of a new pyridinofullerene ligand capable of forming axially symmetric complexes with metalloporphyrins is reported. X-ray structural and molecular modeling studies, (1)H NMR, UV-vis spectroscopy, electrochemistry studies, and fluorescence quenching data support the formation of a strong complex between the new ligand and the metal center of ZnTPP. On the basis of computational studies, the highest occupied molecular orbital (HOMO) of this ligand is significantly different from a model compound with insulating carbons between the pyridine and the fullerene. The N-pyridinium fulleropyrrolidine salts of the new ligand and model compound were also prepared and their spectral and electrochemical properties are reported.

Detection of oligonucleotide hybridization at femtomolar level and sequence-specific gene analysis of the Arabidopsis thaliana leaf extract with an ultrasensitive surface plasmon resonance spectrometer.

A flow-injection (FI) device is combined, through the use of a low-volume (4 microl) flow cell, with an ultrasensitive surface plasmon resonance (SPR) spectrometer equipped with a bi-cell photodiode detector. The application of this novel FI-SPR device for sequence-specific ultratrace analysis of oligodeoxynucleotides (ODNs) and polydeoxynucleotides was demonstrated. Self-assembled monolayers of ODN probes are tethered onto Au films with a mercaptohexyl group at the 3' ends. The FI-SPR provides a detection level (< or =54 fM) 2-3 orders of magnitude lower than other SPR devices and compares well with several ultrasensitive detection methods for labeled DNA targets (e.g. fluorophore-tagged and radiolabeled DNA samples). The technique is also highly selective, since a 47mer ODN target with a single-base mismatch yielded a much smaller SPR signal, and a specific interaction was detected when the complementary target was present at 0.001% of the total DNA. The FI-SPR was extended to the measurement of two individual genes in a cDNA mixture transcribed from an Arabidopsis thaliana leaf mRNA pool. The greatly enhanced sensitivity not only obviates the necessity of DNA labeling, but also significantly reduces sample consumption, allowing direct quantification of low abundance mRNAs in cellular samples without amplification.

All-solid-state carbonate-selective electrode based on a molecular tweezer-type neutral carrier with solvent-soluble conducting polymer solid contact.

Carbonate-selective membranes were prepared by incorporating a molecular tweezer-type carbonate-selective neutral carrier [N,N-dioctyl-3alpha,12alpha-bis(4-trifluoroacetylbenzyloxy)-5beta-cholan-24-amide] into a room temperature vulcanizing-type silicone rubber (3140 RTV-SR) matrix, and deposited on the planar-type electrodes (Pt containing Ag/AgCl electrodes formed on a ceramic plate) with and without an intermediary conducting polymer layer. Two types of solvent-soluble conducting polymers [poly(1-hexyl-3,4-dimethyl-2,5-pyrrolylene) or poly(3-octylthiophene-2,5-diyl)] have been examined as the solid contact material. Potentiometric properties of the resultant all-solid-state electrodes were evaluated in terms of their carbonate selectivity, response slope, potential stability and reproducibility. The sensitivity and carbonate selectivity of the SR membrane-based all-solid-state electrodes with conducting polymer solid contact were comparable to those of conventional electrodes. Experimental results also showed that the intermediary conducting polymer layer used in the all-solid-state electrodes greatly reduces the interference from dissolved oxygen.