Zeev Rosenzweig

Gold Nanoparticle−Quantum Dot−Polystyrene Microspheres as Fluorescence Resonance Energy Transfer Probes for Bioassays

The paper describes the development of highly sensitive particle-based fluorescence resonance energy transfer (FRET) probes that do not use molecular fluorophores as donors and acceptors. In these probes, CdSe/ZnS luminescent quantum dots (QDs) were capped with multiple histidine-containing peptides to increase their aqueous solubility while maintaining their high emission quantum yield and spectral properties. The peptide-modified QDs (QD-His) were covalently attached to carboxyl-modified polystyrene (PS) microspheres to form highly emitting PS microspheres (QD-PS). Gold nanoparticles (AuNPs) were then covalently attached to the QD-PS surface to form AuNP-QD-PS composite microspheres that were used as FRET probes. Attachment of AuNPs to QD-PS completely quenched the QD emission through FRET interactions. The emission of QD-PS was restored when the AuNPs were removed from the surface by thiol ligand displacement. The new AuNP-QD-PS FRET platform is simple to prepare and highly stable, and it opens many new possibilities for carrying out FRET assays on microparticle-based platforms and in microarrays. The versatility of these assays could be greatly increased by replacing the linkers between the QDs and AuNPs with ones that selectively respond to specific cleaving agents or enzymes.

Fiber-optic oxygen sensor based on the fluorescence quenching of tris (5-acrylamido, 1,10 phenanthroline) ruthenium chloride

Abstract Leaking of immobilized fluorescent indicators from the sensing matrix affect the stability of optochemical sensors and, therefore, presents a major problem in this field. When a polymer is used as a matrix support, a covalent bonding of the fluorescence indicator to the matrix eliminates the leaking problem. However, covalent bonding of the indicator to the support cannot be used as a general approach since it may modify the spectral properties of the indicator and its sensing response toward the analyte. This paper describes the synthesis of a modified ruthenium phenanthroline complex which is suitable for acryl co-polymerization with acrylamide. The newly synthesized dye, tris (5 acrylamido, 1,10 phenanthroline) ruthenium chloride, Ru(5-acrylamido-phen)3, maintains its full oxygen response with a slight decrease in its quantum yield for emission. Leaking measurements show a significant improvement in the stability of fiber-optic oxygen sensors that use Ru(5-acrylamido-phen)3 as an oxygen sensitive fluorescence indicator, over previous fiber-optic oxygen sensors that make use of other ruthenium diimine complexes for oxygen measurement. The improvement in stability is particularly noticeable in samples of increased hydrophobicity.

Quantum Dot FRET-Based Probes in Thin Films Grown in Microfluidic Channels

This paper describes the development of new fluorescence resonance energy transfer (FRET)-based quantum dot probes for proteolytic activity. The CdSe/ZnS quantum dots are incorporated into a thin polymeric film, which is prepared by layer-by-layer deposition of alternately charged polyelectrolytes. The quantum dots, which serve as fluorescent donors, are separated from rhodamine acceptor molecules, which are covalently attached to the film surface by a varying number of polyelectrolyte layers. When excited with visible light, the emission color of the polyelectrolyte multilayer film appears orange due to FRET between the quantum dots and molecular acceptors. The emission color changes to green when the rhodamine molecules are removed from the surface by enzymatic cleavage. The new probe design enables the use of quantum dots in bioassays, in this study for real-time monitoring of trypsin activity, while alleviating concerns about their potential toxicity. Application of these quantum dot FRET-based probes in microfluidic channels enables bioanalysis of volume-limited samples and single-cell studies in an in vivo-like environment.

Quantum dot–NBD–liposome luminescent probes for monitoring phospholipase A2 activity

In this paper we describe the fabrication and characterization of new liposome encapsulated quantum dot–fluorescence resonance energy transfer (FRET)-based probes for monitoring the enzymatic activity of phospholipase A2. To fabricate the probes, luminescent CdSe/ZnS quantum dots capped with trioctylphosphine oxide (TOPO) ligands were incorporated into the lipid bilayer of unilamellar liposomes with an average diameter of approximately 100xa0nm. Incorporating TOPO capped quantum dots in liposomes enabled their use in aqueous solution while maintaining their hydrophobicity and excellent photophysical properties. The phospholipid bilayer was labeled with the fluorophore NBD C6-HPC (2-(6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoyl-1-hexa decanoyl-sn-glycero-3-phosphocholine). The luminescent quantum dots acted as FRET donors and the NBD dye molecules acted as FRET acceptors. The probe response was based on FRET interactions between the quantum dots and the NBD dye molecules. The NBD dye molecules were cleaved and released to the solution in the presence of the enzyme phospholipase A2. This led to an increase of the luminescence of the quantum dots and to a corresponding decrease in the fluorescence of the NBD molecules, because of a decrease in FRET efficiency between the quantum dots and the NBD dye molecules. Because the quantum dots were not attached covalently to the phospholipids, they did not hinder the enzyme activity as a result of steric effects. The probes were able to detect amounts of phospholipase A2 as low as 0.0075xa0Uxa0mL−1 and to monitor enzyme activity in real time. The probes were also used to screen phospholipase A2 inhibitors. For example, we found that the inhibition efficiency of MJ33 (1-hexadecyl-3-(trifluoroethyl)-sn-glycero-2-phosphomethanol) was higher than that of OBAA (3-(4-octadecyl)benzoylacrylic acid).

Addition of Fluorescence Lifetime Spectroscopy to the Tool Kit Used to Study the Formation and Degradation of Luminescent Quantum Dots in Solution

The increasing commercialization of consumer electronic products that make use of II-VI semiconductor quantum dots (QDs) has raised significant concerns about their impact on natural systems and human health once they are released into the environment at the end of the products lifetime. In this paper, we demonstrate the addition of fluorescence lifetime spectroscopy to the existing tool kit of spectroscopic techniques to quantitatively monitor changes in QD properties as they form and degrade in solution. Our study reveals that because of its rich information content, fluorescence lifetime spectroscopy has a limited utility as a stand-alone technique in the study of QD formation and degradation. However, combining fluorescence lifetime spectroscopy with the commonly used emission quantum yield and peak width measurements along with other analytical methods, including ultraviolet-visible spectroscopy, transmission electron microscopy, and inductively coupled plasma mass spectrometry measurements, significantly enhances the existing analytical tool kit and provides the capability to monitor in real time, the formation and degradation of luminescent QDs in organic and aqueous solutions under environmentally relevant conditions.

Dynamic analytical chemistry: A kinetic study of the labeling of normal and age fractionated human erythrocytes with monobromobimane

Abstract The application of digital fluorescence imaging microscopy for the analysis of the thiol content and labeling kinetics of normal and age fractionated erythrocytes on a single cell basis is described. Monobromobimane (mBBr), a thiol selective dye, reacts primarily with glutathione, the major thiol compound in erythrocytes. The fluorescence intensity of thiol bound mBBr molecules is about 14-fold higher than the fluorescence intensity of unbound mBBr molecules. The labeling process is monitored in-situ using a slow scan high performance charge coupled device (CCD) camera. A 4-fold cell-to-cell variation is found in the maximum fluorescence intensity of mBBr labeled cells within a large normal population of erythrocytes. This observation is in agreement with previous results obtained for the same cellular system using capillary electrophoresis and laser excited fluorescence detection. In addition, a 2-fold variation is observed in the rate of labeling of the cells with mBBr. Quantitative kinetic measurements of the labeling of age fractionated erythrocytes with mBBr show that the cellular variation in the labeling rate and in the cellular thiol content are related to the cell age.

Structure–Property Relationships of Amine-rich and Membrane-Disruptive Poly(oxonorbornene)-Coated Gold Nanoparticles

The article describes the interactions between poly (oxonorbornenes) (PONs)-coated gold nanoparticles (AuNPs) with phospholipid vesicles and shows that the strength of these interactions strongly depends on the molecular structure of PONs, specifically their amine/alkyl side chain ratio. PONs, which are a recently introduced class of cationic polyelectrolytes, can be systematically varied to control the amine/alkyl ratio and to explore how the chemical character of cationic polyelectrolytes affects their interactions and the interactions of their nanoparticle conjugates with model membranes. Our study shows that increasing the amine/alkyl ratio by copolymerization of diamine and 1:1 amine/butyl oxonorbornene monomers impacts the availability of PONs amine/ammonium functional groups to interact with phospholipid membranes, the PONs surface coverage on AuNPs, and the membrane disruption activity of free PONs and PONs-AuNPs. The study makes use of transmission electron microscopy, UV-vis spectroscopy, dynamic light scattering, thermogravimetric analysis, fluorescamine assay, ζ-potential measurements, and X-ray photoelectron spectroscopy measurements to characterize the PONs-AuNPs size, size distribution, aggregation state, surface charge, and PONs surface coverage. The study also makes use of real-time fluorescence measurements of fluorescent liposomes before and during exposure to free PONs and PONs-AuNPs to determine the membrane disruption activity of free PONs and PONs-AuNPs. As commonly observed with cationic polyelectrolytes, both free PONs and PONs-AuNPs display significant membrane disruption activity. Under conditions where the amine/alkyl ratio in PONs maximizes PONs surface coverage, the membrane disruption activity of PONs-AuNPs is about 10-fold higher than the membrane disruption activity of the same free PONs. This is attributed to the increased local concentration of ammonium ions when PONs-AuNPs interact with the liposome membranes. In contrast, the hydrophobicity of amine-rich PONs, which are made for example from diamine oxonorbornene monomers, is significantly reduced. This leads to a significant reduction of PON surface coverage on AuNPs and in turn to a significant decrease in membrane disruption.

Adverse Interactions of Luminescent Semiconductor Quantum Dots with Liposomes and Shewanella oneidensis

Cadmium-containing luminescent quantum dots (QD) are increasingly used in display, bioimaging, and energy technologies; however, significant concerns have been raised about their potentially adverse impact on human health and the environment. This study makes use of a broad toolkit of analytical methods to investigate and increase our understanding of the interactions of luminescent cadmium-containing (CdSe) and cadmium-free (ZnSe) QD, with and without a passivating higher bandgap energy ZnS shell, with phospholipid vesicles (liposomes), which model bacterial membranes, and with Shewanella oneidensis MR-1, an environmentally relevant bacteria. A unique feature of this study is that all QD types have the same surface chemistry, being capped with uncharged poly(ethylene glycol) ligands. This enables focusing the study on the impact of the QD core on liposomes and bacterial cells. The study reveals that QD association with liposome and bacterial cell membranes is imperative for their adverse impact on liposomes and bacterial cells. The QD concentration-dependent association with liposomes and bacterial cells destabilizes the membranes mechanically, which leads to membrane disruption and lysis in liposomes and to bacterial cell death. The study also shows that cadmium-containing QD exhibit a higher level of membrane disruption in bacterial cells than cadmium-free QD. ZnSe QD have low membrane impact, and coating them with a ZnS shell decreases their membrane disruption activity. In contrast, CdSe QD exhibit a high level of membrane impact, and coating them with a ZnS shell does not decrease, but in fact further increases, their membrane disruption activity. This behavior might be attributed to higher affinity and association of CdSe/ZnS QD with liposomes and bacterial cells and to a contribution of dissolved zinc ions from the ZnS shell to increased membrane disruption activity.

Synthesis and Degradation of Cadmium-Free InP and InPZn/ZnS Quantum Dots in Solution

This study advances the chemical research community toward the goal of replacing toxic cadmium-containing quantum dots (QDs) with environmentally benign InP QDs. The InP QD synthesis uniquely combines the previously reported use of InP magic-sized clusters (MSCs) as a single-source precursor for indium and phosphorus to form InP QDs, with zinc incorporation and subsequent ZnS shelling, to form InPZn/ZnS QDs with luminescence properties comparable to those of commonly used cadmium-containing luminescent QDs. The resulting InPZn/ZnS QDs have an emission quantum yield of about 50% across a broad range of emission peak wavelengths and emission peaks averaging 50 nm fwhm. The emission peak wavelength can be easily tuned by varying the Zn/In ratio in the reaction mixture. The strategy of using zinc stearate to tune the emission properties is advantageous as it does not lead to a loss of emission quantum yield or emission peak broadening. Although the initial optical properties of InP and InPZn/ZnS QDs are promising, thermal stability measurements of InPZn QDs show significant degradation in the absence of a shell compared to the CdSe QDs particularly at increased temperature in the presence of oxygen, which is indicative of thermal oxidation. There is no significant difference in the degradation rate of InP QDs made from molecular precursors and from MSCs. Additionally, the emission intensity and quantum yield of InPZn/ZnS QDs when purified and diluted in organic solvents under ambient conditions decrease significantly compared to those of CdSe/ZnS QDs. This indicates instability of the ZnS shell when prepared by common literature methods. This must be improved to realize high-quality, robust Cd-free QDs with the capability of replacing CdSe QDs in QD technologies.

Release, detection and toxicity of fragments generated during artificial accelerated weathering of CdSe/ZnS and CdSe quantum dot polymer composites

Next generation displays and lighting applications are increasingly using inorganic quantum dots (QDs) embedded in polymer matrices to impart bright and tunable emission properties. The toxicity of some heavy metals present in commercial QDs (e.g. cadmium) has, however, raised concerns about the potential for QDs embedded in polymer matrices to be released during the manufacture, use, and end-of-life phases of the material. One important potential release scenario that polymer composites can experience in the environment is photochemically induced matrix degradation. This process is not well understood at the molecular level. To study this process, the effect of an artificially accelerated weathering process on QD–polymer nanocomposites has been explored by subjecting CdSe and CdSe/ZnS QDs embedded in poly(methyl methacrylate) (PMMA) to UVC irradiation in aqueous media. Significant matrix degradation of QD–PMMA was observed along with measurable mass loss, yellowing of the nanocomposites, and a loss of QD fluorescence. While ICP-MS identified the release of ions, confocal laser scanning microscopy and dark-field hyperspectral imaging were shown to be effective analytical techniques for revealing that QD-containing polymer fragments were also released into aqueous media due to matrix degradation. Viability experiments, which were conducted with Shewanella oneidensis MR-1, showed a statistically significant decrease in bacterial viability when the bacteria were exposed to highly degraded QD-containing polymer fragments. Results from this study highlight the need to quantify not only the extent of nanoparticle release from a polymer nanocomposite but also to determine the form of the released nanoparticles (e.g. ions or polymer fragments).