Intratumoral Biosynthesis of Gold Nanoclusters by Pancreatic Cancer to Overcome Delivery Barriers to Radiosensitization.

Nanoparticle delivery to solid tumors is a prime challenge in nanomedicine. Here, we approach this challenge through the lens of biogeochemistry, the field that studies the flow of chemical elements within ecosystems as manipulated by living cellular organisms and their environments. We leverage biogeochemistry concepts related to gold cycling against pancreatic cancer, considering mammalian organisms as drivers for gold nanoparticle biosynthesis. Sequestration of gold nanoparticles within tumors has been demonstrated as an effective strategy to enhance radiotherapy; however, the desmoplasia of pancreatic cancer impedes nanoparticle delivery. Our strategy overcomes this barrier by applying an atomic-scale agent, ionic gold, for intratumoral gold nanoparticle biosynthesis. Our comprehensive studies showed the cancer-specific synthesis of gold nanoparticles from externally delivered gold ions in vitro and in a murine pancreatic cancer model in vivo; a substantial colocalization of gold nanoparticles (GNPs) with cancer cell nuclei in vitro and in vivo; a strong radiosensitization effect by the intracellularly synthesized GNPs; a uniform distribution of in situ synthesized GNPs throughout the tumor volume; a nearly 40-day total suppression of tumor growth in animal models of pancreatic cancer treated with a combination of gold ions and radiation that was also associated with a significantly higher median survival versus radiation alone (235 vs 102 days, respectively).

Synthesis of an enediyne carbon-allotrope surface for photo-thermal degradation of DNA.

The improper disposal of hospital waste products containing genetic materials poses a serious safety threat. We present herein an environmentally friendly technology using a graphene-based novel carbon-allotropic surface to remediate such wastes. The used carbon-allotrope is decorated with an enediyne (EDE-1) enriched aromatic pi-conjugated structure to create an efficient and active surface for cleaving DNA strands. Under controlled exposure of ultraviolet (UV) radiation and heat, the developed surface influences genetic degradation without disturbing the bacterial populations present downstream of the water treatment system. The designed material has been extensively characterized using physicochemical and biological tools. Our results indicate that this approach can possibly be introduced in large scale hospital waste disposal streams for remediating genetic hazards and thereby developing a portable self-contained system.

Prospecting Cellular Gold Nanoparticle Biomineralization as a Viable Alternative to Prefabricated Gold Nanoparticles.

Gold nanoparticles (GNPs) have shown considerable potential in a vast number of biomedical applications. However, currently there are no clinically approved injectable GNP formulations. Conversely, gold salts have been used in the clinic for nearly a century. Further, there is evidence of GNP formation in patients treated with gold salts (i.e., chrysiasis). Recent reports evaluating this phenomenon in human cells and in murine models indicate that the use of gold ions for in situ formation of theranostic GNPs could greatly improve the delivery within dense biological tissues, increase efficiency of intracellular gold uptake, and specificity of GNP formation within cancer cells. These attributes in combination with safe clinical application of gold salts make this process a viable strategy for clinical translation. Here, the first summary of the current knowledge related to GNP biomineralization in mammalian cells is provided along with critical assessment of potential biomedical applications of this newly emergent field.

A Simplistic Single-Step Method for Preparing Biomimetic Nanoparticles from Endogenous Biomaterials.

Many works utilize products isolated from nature as capping agents to functionalize gold nanoparticles for targeting and therapeutic applications. Some of the most advanced of these strategies utilize complex multicomponent biomaterials, such as whole cell-membranes, for nanoparticle functionalization strategies for evading or initializing immune response as well as for targeting. Strategies like these, wherein whole cell membrane is utilized for functionalization, take advantage of the complexity of the protein-lipid content and organization, which cells normally use for communication and interaction (instilling these capacities to nanoparticle vectors). Many approaches for achieving this in functionalizing the surface of nanoparticles rely on multistep processes, which necessitate the addition and then removal of synthetic molecules, heating, or pH modifications. These processes can have deleterious modifying effects on the functionalizing biomolecules, resulting in loss of product and time during each purification step, as well as potentially changing the biomolecule functionality toward a nondesirable effect. Here, we describe methods for forming gold nanoparticles at room temperature in a single step, functionalized with proteins, using nicotinamide adenine dinucleotide (NADH). This process enables formation of nanoparticles that can be functionalized by individual proteins (demonstrated with FBS) or whole cells membrane (extracted from B16F10 cells). This work is derivative from observations found in the literature by us and others, that mammalian cells are capable of producing gold nanoparticles from ionic gold without the supplementation of chemical species. The products of this single-step synthesis described herein have been optimized to maintain biomolecule integrity and so that there are no further purification steps required. To characterize the nanoparticles in terms of their shape, size, surface functionality, and biomolecule integrity throughout development, we employed light-based spectroscopy techniques, molecular modeling, electron microscopy, light scattering, and gel electrophoresis techniques. In order to compare the optimized biomolecule-functionalized nanoparticles against current standards (which require synthetic linkers, heating, or pH manipulation), we employed metabolic and live/dead assays as well as light-based microscopy/spectroscopy in vitro. In comparing our synthetic process against others for forming gold nanoparticles functionalized with complex biomolecule components (whole-cell membrane), we found that this process had superior particle internalization. Our strategy has similar outlets for application to these other works, however, because this process is entirely reliant on endogenous biomaterials and has additional potential.

Intratumoral generation of photothermal gold nanoparticles through a vectorized biomineralization of ionic gold.

Various cancer cells have been demonstrated to have the capacity to form plasmonic gold nanoparticles when chloroauric acid is introduced to their cellular microenvironment. But their biomedical applications are limited, particularly considering the millimolar concentrations and longer incubation period of ionic gold. Here, we describe a simplistic method of intracellular biomineralization to produce plasmonic gold nanoparticles at micromolar concentrations within 30 min of application utilizing polyethylene glycol as delivery vector for ionic gold. We have characterized this process for intracellular gold nanoparticle formation, which progressively accumulates proteins as the ionic gold clusters migrate to the nucleus. This nano-vectorized application of ionic gold emphasizes its potential biomedical opportunities while reducing the quantity of ionic gold and required incubation time. To demonstrate its biomedical potential, we further induce in-situ biosynthesis of gold nanoparticles within MCF7 tumor mouse xenografts which is followed by its photothermal remediation.

Complementary Oligonucleotide Conjugated Multicolor Carbon Dots for Intracellular Recognition of Biological Events.

By using complementary DNA sequences as surface ligands, we selectively allow two individual diffusing "dual-color" carbon dots to interact in situ and in vitro. Spontaneous nanoscale oxidation of surface-abundant nitroso-/nitro-functionalities leads to two distinctly colored carbon dots (CD) which are isolated by polarity driven chromatographic separation. Green- and red-emitting carbon dots (gCD and rCD) were decorated by complementary single-stranded DNAs which produce a marked increase in the fluorescence emission of the respective carbon dots. Mutual colloidal interactions are achieved through hybridization of complementary DNA base pairs attached to the respective particles, resulting in quenching of their photoluminescence. The observed post-hybridization quenching is presumably due to a combined effect from an aggregation of CDs post duplex DNA formation and close proximity of multicolored CDs, having overlapped spectral regions leading to a nonradiative energy transfer process possibly released as heat. This strategy may contribute to the rational design of mutually interacting carbon dots for a better control over the resulting assembly structure for studying different biological phenomenon including molecular cytogenetics. One of the newly synthesized CDs was successfully used to image intracellular location of GAPDH mRNA using an event of change in fluorescence intensity (FI) of CDs. This selectivity was introduced by conjugating an oligonucleotide harboring complementary sequence to GAPDH mRNA. FI of this conjugated carbon dot, rCD-GAPDH, was also found to decrease in the presence of Ca2+, varied in relation to H+ concentrations, and could serve as a tool to quantify the intracellular concentrations of Ca2+ and pH value (H+) which can give important information about cell survival. Therefore, CD-oligonucleotide conjugates could serve as efficient probes for cellular events and interventions.

Enzyme-catalysed biodegradation of carbon dots follows sequential oxidation in a time dependent manner.

Carbon dots (CDs) have recently garnered significant attention owing to their excellent luminescence properties, thereby demonstrating a variety of applications in in vitro and in vivo imaging. Understanding the long-term metabolic fate of these agents in a biological environment is the focus of this work. Here we show that the CDs undergo peroxide catalysed degradation in the presence of lipase. Our results indicate that differently charged CD species exhibit unique degradation kinetics upon being subjected to enzyme oxidation. Furthermore, this decomposition correlates with the relative accessibility of the enzymatic molecule. Using multiple physico-chemical characterization studies and molecular modelling, we confirmed the interaction of passivating surface abundant molecules with the enzyme. Finally, we have identified hydroxymethyl furfural as a metabolic by-product of the CDs used here. Our results indicate the possibility and a likely mechanism for complete CD degradation in living systems that can pave the way for a variety of biomedical applications.

Electrochemical-digital immunosensor with enhanced sensitivity for detecting human salivary glucocorticoid hormone.

In this work, an ultra-sensitive electrochemical-digital sensor chip is devised for potential use as a digital stress analyzer for point-of-care testing (POCT) and preventive on-site recording of the hormone 'cortisol', a glucocorticoid class of steroid hormone present in the human saliva. The sensor was interfaced and re-configured with a high precision impedance converter system (AD5933) and used for electrochemical impedance spectroscopy (EIS) to evaluate the cortisol levels in seven saliva samples. To obtain enhanced biological (cortisol) recognition and achieve a lower limit of detection 0.87 ± 0.12 pg mL-1 (2.4 ± 0.38 pmol mL-1) with a wide range from 1 pg mL-1 to 10 ng mL-1 (2.75 pmol mL-1 to 27.58 pmol mL-1; R2 = 0.9831), bovine serum albumin (1% BSA) was utilized as an effective sensitivity enhancer in addition to optimizing the other two parameters: (i) anti-cortisol antibody (anti-CAb) covalently attached to micro-Au electrodes and (ii) saliva sample incubation time on the sensor chip. The results obtained in this work were corroborated with the gold standard ELISA test with an accuracy of 96.3% and other previously reported biosensors. We envisage that the conceivable standpoint of this study can be a practice towards new development in cortisol biosensing, which will be pertinent to POCT targeted for in vitro psychobiological study on patient cortisol in saliva, and finally an implantable sensor chip in the future.

Carbon dots with induced surface oxidation permits imaging at single-particle level for intracellular studies.

For robust single particle optical detection, a high sensitivity in photoluminescence (PL) of Carbon Dots (CDs) must be achieved. PL sensitivity can be successfully correlated with their surface chemistry but requires high synthetic control without altering their basic surface properties. Here we describe conditions for the controlled synthesis of CDs that resulted in a PL sensitivity at the single-particle level. We report that a stoichiometric catalyst N-methyl morpholine-N-oxide (NMMO) can be used as a 'sacrificial' single additive to aid nanoscale surface oxidation. A 24 h NMMO-mediated oxidation increased coverage of oxidized nanoscale surface 3% to 20.9%. NMMO-oxidized CDs (CD-NMMOs) display superior particle brightness, as evidenced by the increase of light absorbance and an enhancement of quantum yield which is characterized by a series of physicochemical and biophysical experiments. We also demonstrate that CD-NMMOs is well suited for intracellular and single-particle imaging.

In situ plasmonic generation in functional ionic-gold-nanogel scaffold for rapid quantitative bio-sensing.

Conventional analytical techniques, which have been developed for high sensitivity and selectivity for the detection and quantification of relevant biomarkers, may not be as suitable for medical diagnosis in resource scarce environments as compared to point-of-care devices (POC). We have developed a new reactive sensing material which contains ionic gold entrapped within an agarose gel scaffold for POC quantification of ascorbic acid (AA) in tear fluid. Pathologically elevated concentration of AA in human tear fluid can serve as a biomarker for full-thickness injuries to the ocular surface, which are a medical emergency. This reactive sensing material will undergo colorimetric changes, quantitatively dependent on endogenous bio-reductants that are applied, as the entrapped ionic gold is reduced to form plasmonic nanoparticles. The capacity for this reactive material to function as a plasmonically driven biosensor, called 'OjoGel' (ojo-eye), was demonstrated with the endogenous reducing agent, AA. Through applications of AA of varied concentrations to the OjoGel, we demonstrated a quantitative colorimetric relationship between red (R) hexadecimal values and concentrations of AA in said treatments. This colorimetric relationship is directly resultant of plasmonic gold nanoparticle formation within the OjoGel scaffold. Using a commercially available mobile phone-based Pixel Picker® application, the OjoGel plasmonic sensing platform opens a new avenue for easy-to-use, rapid, and quantitative biosensing with low cost and accurate results.

Electrically-receptive and thermally-responsive paper-based sensor chip for rapid detection of bacterial cells.

Although significant technological advancements have been made in the development of analytical biosensor chips for detecting bacterial strains (E. coli, S. Mutans and B. Subtilis), critical requirements i.e. limit of detection (LOD), fast time of response, ultra-sensitivity with high reproducibility and good shelf-life with robust sensing capability have yet to be met within a single sensor chip. In order to achieve these criteria, we present an electrically-receptive thermally-responsive (ER-TR) sensor chip comprised of simple filter paper used as substrate coated with composite of poly(N-isopropylacrylamide) polymer (PNIPAm) - graphene nanoplatelet (GR) followed by evaporation of Au electrodes for capturing both Gram-positive (S. mutans and B. subtilis) and Gram-negative (E. coli) bacterial cells in real-time. Autoclave water, tap water, lake water and milk samples were tested with ER-TR chip with and without bacterial strains at varying concentration range 101-105 cells/mL. The sensor was integrated with in-house built printed circuit board (PCB) to transmit/receive electrical signals. The interaction of E. coli, S. mutans and B. subtilis cells with fibers of PNIPAm-GR resulted in a change of electrical resistance and the readout was monitored wirelessly in real-time using MATLAB algorithm. Finally, prepared ER-TR chip exhibited the reproducibility of 85-97% with shelf-life of up to four weeks after testing with lake water sample.

Facile Chemical Strategy to Hydrophobically Modify Solid Nanoparticles Using Inverted Micelle-Based Multicapsule for Efficient Intracellular Delivery.

Theranostic nanoparticles have incredible potential for biomedical applications by enabling visual confirmation of therapeutic efficacy. Numerous issues challenge their clinical translation and are primarily related to the complex chemistry and scalability of synthesizing Nanoparticles. We report a 2-step chemical strategy for high-throughput intracellular delivery of organic and inorganic solid nanoparticles. This process takes an additional step beyond hydrophobic surface modification facilitated by inverted micelle transfer, toward the packing of multiple solid nanoparticles into a soft-shelled lipid capsule, termed the Nano-multicapsule (NMC). This technique is high yielding and does not require the complex purification steps in anaerobic/hydrophobic reactions for hydrophobic modification. To demonstrate the efficacy across different material compositions, we separately entrapped ∼10 nm gold and carbon nanoparticles (AuNP and CNP) within inverted micelles, and subsequently NMCs, then quantified their internalization in a human breast cancer cell line. For encapsulated AuNPs (NMC-AuNP), we confirmed greater cellular internalization of gold through ICP-OES and TEM analyses. Raman spectroscopic analysis of cells treated with encapsulated CNPs (NMC-CNP) also exhibited high degrees of uptake with apparent intracellular localization as opposed to free CNP treatment.

α-Amino Acid Rich Photophytonic Nanoparticles of Algal Origin Serendipitously Reveal Antimigratory Property against Cancer.

Spheroidal nanoparticles of algal ("phytonic") origin were synthesized and composed of carbonaceous architectures and surface-rich oxygenated functional groups. Nanoparticles were negatively charged and efficiently luminescent after ultraviolet-range excitation and called as "photophytonic" nanoparticles. A multitude of analytical techniques confirmed the rich profusion of hydroxyl, carboxylate, and amines at the nanoscale, while spectroscopic investigation indicated the presence of α-amines, a signature functionality present in amino acids. Confirmed via a series of biological assays, i.e., growth regression, antimigration, and protein-regression studies, photophytonic nanoparticles serendipitously revealed remarkable anticancer activity against various stages of breast cancer cells, barring the need for an encapsulated drug. We report that nanoparticles derived from algal biomass exhibit intrinsic antimigratory properties against cancer, likely due to the rich abundance of α-amino acids.

Genomic DNA Interactions Mechanize Peptidotoxin-Mediated Anticancer Nanotherapy.

Host defense peptides (HDPs) are a class of evolutionarily conserved substances of the innate immune response that have been identified as major players in the defense system in many living organisms. Some of the HDPs are also referred to as peptidotoxins, which offer immense potential for anticancer therapy. However, their therapeutic potential is yet to be fully translated mainly due to their off-target toxicity. Here we show that their nanoenabled delivery may become beneficial in controlling their delivery in intracellular space. We introduced an amphiphilic polymer to synthesize a well-defined, self-assembled, rigid-cored polymeric nanoarchitecture for controlled delivery of three model peptidotoxins, i.e., melittin, TSAP-1, and a negative control peptide of synthetic origin. Interestingly, our results revealed strong interaction of peptidotoxins with duplex plasmid DNA. Extensive biophysical characterization (UV-vis spectroscopy, gel electrophoresis, MTT assay, and flow assisted cell sorting) experimentally verified that peptidotoxins were able to interact with genomic DNA in vitro and in turn influence the cancer cell growth. Thus, we unraveled that, through genomic DNA regulation, peptidotoxins can play a role in cell cycle regulation and exert their anticancer activities.

Nanosalina: A Tale of Saline-Loving Algae from the Lake's Agony to Cancer Therapy.

The nanoparticles (NPs) that contain the therapeutic agent within themselves without further modifications can be coined as "self-therapeutic" NPs. The development of these agents especially when derived from natural resources can lead to a paradigm shift in the field of cancer nanotechnology as they can immensely facilitate the complex chemistry procedures and the follow up biological complications. Herein, we demonstrate that inherently therapeutic NPs "integrating" ß-carotene can be synthesized from Dunaliella salina microalgae in a single step without complicated chemistry. The facile synthesis involved microwave irradiation of aqueous suspension of algae which resulted in water dispersible NPs with hydrodynamic diameter of ∼80 nm. Subsequently, extensive physiochemical characterizations were performed to confirm the integrity of the particles. The pro-oxidant activities of the integrated ß-carotene were triggered by photoexcitation under UV lamp (362 nm). It was demonstrated that after UV exposure, the C32 human melanoma cells incubated with NPs experienced extensive cell death as opposed to nonilluminated samples. Further cellular analysis revealed that the significant reactive oxygen species (ROS) and in particular singlet oxygen were responsible for the cells' damage while the mode of cell death was dominated by apoptosis. Moreover, detailed endocytic inhibition studies specified that UV exposure affected NPs' cellular uptake mechanism. These inherently therapeutic NPs can open new avenues for melanoma cancer treatment via ROS generation in vitro.

Real-Time Monitoring of Post-Surgical and Post-Traumatic Eye Injuries Using Multilayered Electrical Biosensor Chip.

Lack of current techniques for the early monitoring of bleb leaks and other post-traumatic or post-surgical ocular injury has posed an unmet clinical need for the development of new techniques. Present evaluation techniques use either subjective or nonquantitative approaches. At present, there are no FDA approved ocular devices that can directly measure ascorbic acid (AA) concentration levels for both tear film (TF) and aqueous humor (AH) at point-of-care (POC) level. Toward this aim, we present a novel POC quantitative assay, called the ocular biosensor device, which can be used to evaluate the integrity of the anterior surface of the eye by measuring the concentration of AA in TF and AH. Herein, we utilize a novel scientific engineering approach for the development of a disposable paper based POC ocular biosensor strip. A grafted poly(styrene)-block-poly(acrylic acid) (PS-b-PAA) and graphene platelet composite with contour based µ-electrodes design (CBµE) exhibit a highly sensitive platform to perform electrochemical immunosensing technique to study clinical samples that have small volumes like tear fluid. Samples used in this study were collected clinically from subjects undergoing therapeutic anterior chamber paracentesis. The proposed biosensor reports the level of AA concentration on an electronic screen, making the results easy to read, efficient, and reliable.

Paper-Based Analytical Biosensor Chip Designed from Graphene-Nanoplatelet-Amphiphilic-diblock-co-Polymer Composite for Cortisol Detection in Human Saliva.

Cortisol has been identified as a biomarker in saliva to monitor psychological stress. In this work, we report a label-free paper-based electrical biosensor chip to quantify salivary cortisol at a point-of-care (POC) level. A high specificity of the sensor chip to detect cortisol with a detection limit of 3 pg/mL was achieved by conjugating anticortisol antibody (anti-CAB) on top of gold (Au) microelectrodes using 3,3'-dithiodipropionic acid di(N-hydroxysuccinimide ester (DTSP) as a self-assembled monolayer (SAM) agent. The electrode design utilized poly(styrene)-block-poly(acrylic acid) (PS67-b-PAA27) polymer and graphene nanoplatelets (GP) suspension coated on filter paper to increase the sensitivity of the immune response. A biosensor chip was then integrated with a lab-built low-cost miniaturized printed circuit board (PCB) to provide an electrical connection and to wirelessly transmit/receive electrical signals using MATLAB. This fully integrated proposed hand-held device successfully exhibited a wide cortisol-detection range from 3 pg/mL to 10 µg/mL, with a sensitivity of 50 Ω (pg mL-1)-1. The performance of the proposed cortisol sensor chip was validated using an enzyme-linked immunosorbent assay (ELISA) technique with a regression value of 0.9951. The advantages of the newly developed cortisol immune biosensor over previously reported chips include an improved limit of detection, no need for additional redox medium for electron exchange, faster response to achieve stable data, excellent shelf life, and its economical production.

Nano-Cesium for Anti-Cancer Properties: An Investigation into Cesium Induced Metabolic Interference.

The use of cesium chloride (CsCl) for cancer therapy ("high pH therapy") has been theorized to produce anticancer properties by raising intracellular pH to induce apoptosis. Although considered as "alternative medicine", little scientific evidence supports this theory. Alternatively, cells have no cesium ion (Cs+) mediated channels for clearance. Thus, such unstable electrochemical distributions have the severe potential to disrupt electrochemical dependent cellular processes, such as glucose cotransporters. Hence, a detailed investigation of pH changing effects and glucose uptake inhibition are warranted as a possible cesium-induced anticancer therapy. We developed and characterized cesium nanoparticles (38 ± 6 nm), termed NanoCs, for nanoparticle-mediated internalization of the ion, and compared its treatment to free CsCl. Our investigations suggest that neither NanoCs nor CsCl drastically changed the intracellular pH, negating the theory. Alternatively, NanoCs lead to a significant decrease in glucose uptake when compared to free CsCl, suggesting cesium inhibited glucose uptake. An apoptosis assay of observed cell death affirms that NanoCs leads tumor cells to initiate apoptosis rather than follow necrotic behavior. Furthermore, NanoCs lead to in vivo tumor regression, where H&E analysis confirmed apoptotic cell populations. Thus, NanoCs performed pH-independent anticancer therapy by inducing metabolic stasis.

(-)/(+)-Sparteine induced chirally-active carbon nanoparticles for enantioselective separation of racemic mixtures.

Chiral carbon nanoparticles (CCNPs) were developed by surface passivation using the chiral ligand (-)-sparteine or (+)-sparteine (denoted (-)-SP/CNP and (+)-SP/CNP, respectively). The chirality of the prepared CCNPs was demonstrated by circular dichroism and polarimetry and employed as an enantioselective separation platform for representative racemic mixtures.

Dual-modality, fluorescent, PLGA encapsulated bismuth nanoparticles for molecular and cellular fluorescence imaging and computed tomography.

Reports of molecular and cellular imaging using computed tomography (CT) are rapidly increasing. Many of these reports use gold nanoparticles. Bismuth has similar CT contrast properties to gold while being approximately 1000-fold less expensive. Herein we report the design, fabrication, characterization, and CT and fluorescence imaging properties of a novel, dual modality, fluorescent, polymer encapsulated bismuth nanoparticle construct for computed tomography and fluorescence imaging. We also report on cellular internalization and preliminary in vitro and in vivo toxicity effects of these constructs. 40 nm bismuth(0) nanocrystals were synthesized and encapsulated within 120 nm Poly(dl-lactic-co-glycolic acid) (PLGA) nanoparticles by oil-in-water emulsion methodologies. Coumarin-6 was co-encapsulated to impart fluorescence. High encapsulation efficiency was achieved ∼70% bismuth w/w. Particles were shown to internalize within cells following incubation in culture. Bismuth nanocrystals and PLGA encapsulated bismuth nanoparticles exhibited >90% and >70% degradation, respectively, within 24 hours in acidic, lysosomal environment mimicking media and both remained nearly 100% stable in cytosolic/extracellular fluid mimicking media. µCT and clinical CT imaging was performed at multiple X-ray tube voltages to measure concentration dependent attenuation rates as well as to establish the ability to detect the nanoparticles in an ex vivo biological sample. Dual fluorescence and CT imaging is demonstrated as well. In vivo toxicity studies in rats revealed neither clinically apparent side effects nor major alterations in serum chemistry and hematology parameters. Calculations on minimal detection requirements for in vivo targeted imaging using these nanoparticles are presented. Indeed, our results indicate that these nanoparticles may serve as a platform for sensitive and specific targeted molecular CT and fluorescence imaging.