Imaging Viral Behavior in Mammalian Cells with Self‐Assembled Capsid–Quantum‐Dot Hybrid Particles

Unique spectral properties of quantum dots (QDs) enable ultrasensitive and long‐term biolabeling. Aiming to trace the infection, movement, and localization of viruses in living cells, QD‐containing virus‐like particles (VLPs) of simian virus 40 (SV40), termed SVLP‐QDs, are constructed by in vitro self‐assembly of the major capsid protein of SV40. SVLP‐QDs show homogeneity in size (≈24 nm), similarity in spectral properties to unencapsidated QDs, and considerable stability. When incubated with living cells, SVLP‐QDs are shown to enter the cells by caveolar endocytosis, travel along the microtubules, and accumulate in the endoplasmic reticulum. This process mimics the early infection steps of SV40. This is the first paradigm of imaging viral behaviors with encapsidated QDs in living cells. The method may provide a new alternative for various purposes, such as tracing viruses or viral components, targeted nanoparticle delivery, and probing of drug delivery.     

Viral Coat Proteins as Flexible Nano‐Building‐Blocks for Nanoparticle Encapsulation

Viral capsid–nanoparticle hybrid structures offer new opportunities for nanobiotechnology. We previously generated virus‐based nanoparticles (VNPs) of simian virus 40 (SV40) containing quantum dots (QDs) for cellular imaging. However, as an interesting issue of nano‐bio interfaces, the mechanism of nanoparticle (NP) encapsulation by viral coat proteins remains unclear. Here, four kinds of QDs with the same core/shell but different surface coatings are tested for encapsulation. All the QDs can be encapsulated efficiently and there is no correlation between the encapsulation efficiency and the surface charge of the QDs. All the SV40 VNPs encapsulating differently modified QDs show similar structures, fluorescence properties, and activity in entering living cells. These results demonstrate the flexibility of SV40 major capsid protein VP1 in NP encapsulation and provide new clues to the mechanism of NP packaging by viral shells.     

一种简单的水热法制备油溶性PbSe量子点

本研究采用简单的水热法得到了单一形态学和小尺寸分布的油溶性PbSe量子点。所得到的立方相PbSe量子点颗粒呈现近球形, 且平均颗粒尺寸为4.0 ± 0.5 nm。PbSe量子点在435 nm近紫外区域呈现出较强的和相对较窄的光致发光光谱, 光谱的半高宽值约为80 nm。随着反应时间的延长和反应温度的升高, 发光光谱向低能量区域移动, 谱峰的半高宽也随之变大。在改变前驱体Pb/S摩尔比的条件下, 发光光谱相对强度降低, 光谱也发生向长波长区域移动。另外, 随着反应温度和前驱体Pb/S摩尔比的改变, PbSe量子点颗粒表面的缺陷也增多。在反应过程中, 小颗粒长大成大尺寸颗粒促使PbSe量子点的发光光谱向长波长移动, 这个现象符合奥斯瓦尔德熟化定律。热力学不稳定和颗粒表面低浓度油酸包覆也造成PbSe量子点颗粒表面产生缺陷。     

Advanced optical imaging reveals the dependence of particle geometry on interactions between CdSe quantum dots and immune cells

The biocompatibility and possible toxicological consequences of engineered nanomaterials, including quantum dots (QDs) due to their unique suitability for biomedical applications, remain intense areas of interest. We utilized advanced imaging approaches to characterize the interactions of CdSe QDs of various sizes and shapes with live immune cells. Particle diffusion and partitioning within the plasma membrane, cellular uptake kinetics, and sorting of particles into lysosomes were all independantly characterized. Using high-speed total internal reflectance fluorescence (TIRF) microscopy, we show that QDs with an average aspect ratio of 2.0 (i.e., rod-shaped) diffuse nearly an order of magnitude slower in the plasma membrane than more spherical particles with aspect ratios of 1.2 and 1.6, respectively. Moreover, more rod-shaped QDs were shown to be internalized into the cell 2-3 fold more slowly. Hyperspectral confocal fluorescence microscopy demonstrates that QDs tend to partition within the cell membrane into regions containing a single particle type. Furthermore, data examining QD sorting mechanisms indicate that endocytosis and lysosomal sorting increases with particle size. Together, these observations suggest that both size and aspect ratio of a nanoparticle are important characteristics that significantly impact interactions with the plasma membrane, uptake into the cell, and localization within intracellular vesicles. Thus, rather than simply characterizing nanoparticle uptake into cells, we show that utilization of advanced imaging approaches permits a more nuanced and complete examination of the multiple aspects of cell-nanoparticle interactions that can ultimately aid understanding possible mechanisms of toxicity, resulting in safer nanomaterial designs.     

Optical encoding of microbeads based on silica particle encapsulated quantum dots and its applications

A novel method concerning the coding technology of polystyrene beads with Si encapsulated quantum dot (QD) particles (Si@QDs particles) is studied in this paper. In the reverse microemulsion system containing tetraethoxysilane (TEOS), water-soluble QDs (emission peak at 600?nm) were enveloped within the silica shell, forming Si@QDs particles. The Si@QDs particles were characterized by TEM, showing good uniform size, with an average diameter of about 167.0?nm. In comparison with the pure water-soluble QDs, the encapsulation of water-soluble QDs in the silica shell led to an enhancement in anti-photobleaching by providing inert barriers for the QDs. Images presented by SEM and confocal laser scanning microscopy demonstrated that the Si@QDs particles were equably coated on the surface of carboxyl functionalized polystyrene (PS) beads. Then, with the assistance of ethyl-3-(dimethyl aminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS), human IgG could be successfully crosslinked to Si@QDs particle coated PS-COOH beads. Furthermore, the Si@QDs coated PS-COOH beads with human IgG were examined in immunoassay experiments, and the results indicated that these beads could be applied in the specific recognition of goat-anti-human IgG in solution. This investigation is expected to provide a new route to bead coding in the field of suspension microarrays, based on the use of QDs.     

Nanotoxicology: Advanced Optical Imaging Reveals the Dependence of Particle Geometry on Interactions Between CdSe Quantum Dots and Immune Cells (Small 3/2011)

The biocompatibility and possible toxicological consequences of engineered nanomaterials, including quantum dots (QDs) due to their unique suitability for biomedical applications, remain intense areas of interest. We utilized advanced imaging approaches to characterize the interactions of CdSe QDs of various sizes and shapes with live immune cells. Particle diffusion and partitioning within the plasma membrane, cellular uptake kinetics, and sorting of particles into lysosomes were all independantly characterized. Using high‐speed total internal reflectance fluorescence (TIRF) microscopy, we show that QDs with an average aspect ratio of 2.0 (i.e., rod‐shaped) diffuse nearly an order of magnitude slower in the plasma membrane than more spherical particles with aspect ratios of 1.2 and 1.6, respectively. Moreover, more rod‐shaped QDs were shown to be internalized into the cell 2‐3 fold more slowly. Hyperspectral confocal fluorescence microscopy demonstrates that QDs tend to partition within the cell membrane into regions containing a single particle type. Furthermore, data examining QD sorting mechanisms indicate that endocytosis and lysosomal sorting increases with particle size. Together, these observations suggest that both size and aspect ratio of a nanoparticle are important characteristics that significantly impact interactions with the plasma membrane, uptake into the cell, and localization within intracellular vesicles. Thus, rather than simply characterizing nanoparticle uptake into cells, we show that utilization of advanced imaging approaches permits a more nuanced and complete examination of the multiple aspects of cell‐nanoparticle interactions that can ultimately aid understanding possible mechanisms of toxicity, resulting in safer nanomaterial designs.     

Particle Size,Surface Coating,and PEGylation Influence the Biodistribution of Quantum Dots in Living Mice

This study evaluates the influence of particle size, PEGylation, and surface coating on the quantitative biodistribution of near‐infrared‐emitting quantum dots (QDs) in mice. Polymer‐ or peptide‐coated ⁶⁴Cu‐labeled QDs 2 or 12 nm in diameter, with or without polyethylene glycol (PEG) of molecular weight 2000, are studied by serial micropositron emission tomography imaging and region‐of‐interest analysis, as well as transmission electron microscopy and inductively coupled plasma mass spectrometry. PEGylation and peptide coating slow QD uptake into the organs of the reticuloendothelial system (RES), liver and spleen, by a factor of 6–9 and 2–3, respectively. Small particles are in part renally excreted. Peptide‐coated particles are cleared from liver faster than physical decay alone would suggest. Renal excretion of small QDs and slowing of RES clearance by PEGylation or peptide surface coating are encouraging steps toward the use of modified QDs for imaging living subjects.     

Progression of respiratory syncytial virus infection monitored by fluorescent quantum dot probes

We report the use of quantum dots (QDs) to identify the presence and monitor the progression of respiratory syncytial virus (RSV) infection over time by labeling the F and G proteins. In addition, co-localization of these viral proteins was shown using confocal microscopy. The implications of these results are that QDs may provide a method for early, rapid detection of viral infection and open the door for future studies of the intricate spatial features cell trafficking of viral proteins.     

Self-surface passivation of CdX (X = Se, Te) quantum dots

A small portion of a reaction mixture including unpurified CdX (X = Se or Te) quantum dots (QDs), in which unreacted Cd and Se ions were left together with coordinating solvents, was dropped into an organic solvent. The CdX QDs in this organic solution showed enhancement of photoluminescence (PL) efficiency, growth of particles, and focusing of size distribution for more than 10 h at room temperature (RT, -23 degrees C). These effects were attributed to passivation of QDs' surface by Cd and X ions present in the solution. No external energy source was used for these achievements; therefore, the process is termed as self-surface passivation. The self-surface passivation was reproduced using purified CdX QDs with additional Cd and X ions in an organic solvent. The self-surface passivation method was applied to RT-synthesized CdSe QDs, which is characterized by a broad PL spectrum (fwhm - 150 nm) for monodispersed QDs, to modify their emission characteristics. On self-surface passivation, the broad PL spectrum was narrowed (fwhm - 35 nm) and the QDs were grown. The X-ray diffraction measurements of RT-synthesized CdSe QDs and that subsequently aged in 1-butanol showed that crystallinity of the samples was improved on aging.     

Large-scale preparation of CdS quantum dots by direct thermolysis of a single-source precursor

CdS quantum dots (QDs) have been synthesized on a large scale, based on the direct thermolysis of one single-source precursor, (Me(4)N)(4)[S(4)Cd(10)(SPh)(16)], in hexadecylamine (HDA). Transmission electron microscopy (TEM) observations show that the CdS QDs are well-defined, nearly spherical particles. The clear lattice fringes in high-resolution TEM (HRTEM) images confirm the crystalline nature of the QDs. The broad diffraction in the x-ray diffraction (XRD) pattern and diffuse diffraction rings of the selected-area electron diffraction (SAED) pattern are typical of nanomeric-size particles and indicative of the hexagonal phase of CdS QDs. The absorption spectra confirm quantum confinement of CdS QDs. The synthesis process for CdS QDs was investigated by ultraviolet-visible (UV-vis) absorption spectroscopy. The results demonstrate that the nucleation and growth stages were separated automatically in a homogeneous system.     

Fluorescent magnetic hybrid nanoprobe for multimodal bioimaging

A fluorescent magnetic hybrid imaging nanoprobe (HINP) was fabricated by the conjugation of superparamagnetic Fe3O4 nanoparticles and visible light emitting (～600 nm) fluorescent CdTe/CdS quantum dots (QDs). The assembly strategy used the covalent linking of the oxidized dextran shell of magnetic particles to the glutathione ligands of QDs. The synthesized HINP formed stable water-soluble colloidal dispersions. The structure and properties of the particles were characterized by transmission electron and atomic force microscopy, energy dispersive x-ray analysis and inductively coupled plasma optical emission spectroscopy, dynamic light scattering analysis, optical absorption and photoluminescence spectroscopy, and fluorescent imaging. The luminescence imaging region of the nanoprobe was extended to the near-infrared (NIR) (～800 nm) by conjugation of the superparamagnetic nanoparticles with synthesized CdHgTe/CdS QDs. Cadmium, mercury based QDs in HINP can be easily replaced by novel water-soluble glutathione stabilized AgInS2/ZnS QDs to present a new class of cadmium-free multimodal imaging agents. The observed NIR photoluminescence of fluorescent magnetic nanocomposites supports their use for bioimaging. The developed HINP provides dual-imaging channels for simultaneous optical and magnetic resonance imaging.     

Histidine functionalised biocompatible CdS quantum dots synthesised by sonochemical method

Histidine functionalised CdS quantum dots (QDs) have been synthesised by sonochemical method. Transmission Electron Microscopy (TEM) observation shows that the histidine functionalised CdS QDs are well-defined, nearly spherical particles. The X-ray diffraction pattern indicates formation of cubic phase of CdS/histidine QDs. The absorption spectra confirm quantum confinement of histidine functionalised CdS QDs. The photoluminescence property of CdS/histidine QDs is found better than that of CdS QDs. Histidine functionalised CdS QDs, in which histidine acts as a biocompatibiliser, can find potential applications in the biological fields.     

A novel route to synthesize CdS quantum dots on the surface of silk fibers via γ-radiation

CdS quantum dots (QDs) were successfully synthesized on the surface of silk fibers via γ-ray irradiation. Scanning electron microscope (SEM) measurement revealed that the particles on the silk fibers were less than 15nm in diameter. The energy dispersion spectrum (EDS), X-ray diffraction (XRD), laser scanning confocal microscope (LSCM) were also used to identify and investigate the properties of products. Moreover, a mechanism for the formation of CdS QDs on silk fibers under γ-radiation was generally discussed. The resulting silk fibers coated with CdS QDs might be useful as smart structural fabrics in a range of applications.     

Exploring feasibility of multicolored CdTe quantum dots for in vitro and in vivo fluorescent imaging

We report the use of novel multicolored CdTe quantum dots (QDs) as fluorophores for biological fluorescence imaging. The CdTe QDs were prepared to exhibit emission wavelengths in the green, yellow, and red range by using trifluoroacetic acid (TFA), L-cysteine and thioglycolic acid (TGA) as surface stabilizers, respectively. The particles have good water solubility and photostability. Fluorescence imaging potential was evaluated in vitro and in vivo using a multispectral Maestro CRI Fluorescence Imaging system. The results show that different colored CdTe QDs allow sensitive detection simultaneously or separately both in vitro and in vivo against background fluorescence. The studies indicate that CdTe QDs can provide alternative fluorescent probes for biological imaging.     

GFP expression by intracellular gene delivery of GFP-coding fragments using nanocrystal quantum dots

Gene therapy is an attractive approach to supplement a deficient gene function. Although there has been some success with specific gene delivery using various methods including viral vectors and liposomes, most of these methods have a limited efficiency or also carry a risk for oncogenesis. We herein report that quantum dots (QDs) conjugated with nuclear localizing signal peptides (NLSP) successfully introduced gene-fragments with promoter elements, which promoted the expression of the enhanced green fluorescent protein (eGFP) gene in mammalian cells. The expression of eGFP protein was observed when the QD/gene-construct was added to the culture media. The gene-expression efficiency varied depending on multiple factors around QDs, such as (1) the reading direction of the gene-fragments, (2) the quantity of gene-fragments attached on the surface of the QD-constructs, (3) the surface electronic charges varied according to the structure of the QD/gene-constructs, and (4) the particle size of QD/gene complex varied according to the structure and amounts of gene-fragments. Using this QD/gene-construct system, eGFP protein could be detected 28 days after the gene-introduction whereas the fluorescence of QDs had disappeared. This system therefore provides another method for the intracellular delivery of gene-fragments without using either viral vectors or specific liposomes.     

Probing the cytotoxicity of CdSe quantum dots with surface modification

The cytotoxicity of CdSe quantum dots (QDs) with surface modification was reported first in the paper. CdSe QDs were incorporated into poly (d, l) lactide (PLA) nanoparticles and then surface modified with Fluronic® 68 (F-68), cetyltrimethyl ammonium bromide (CTAB) and sodium dodecyl sulfate (SDS), respectively. Three different particle sizes and zeta potential of the surface modified CdSe QDs were produced using a nano-precipitation method. The cytotoxicity of the surface modified CdSe QDs was evaluated in HepG2 cell model with MTT viability assay. The results showed that the cytotoxicity of the surface modified CdSe QDs in vitro was dependent on the surface properties. Surface modification with F-68 and SDS could lessen the cytotoxicity of CdSe QDs, while surface modification with CTAB showed significant cell damage. CdSe QDs surface modified with F-68 were injected into mice and the fluorescence images in viscus were obtained. The results suggested that CdSe QDs surface modified with F-68 have low cytotoxicity and good potential for biological labeling and imaging applications.     

Effect of capping agents on optical and antibacterial properties of cadmium selenide quantum dots

Cadmium selenide quantum dots (CdSe QDs) were synthesized in aqueous phase by the freezing temperature injection technique using different capping agents (viz. thioglycolic acid, 1-thioglycerol, L-cysteine). Absorption spectra of CdSe QDs exhibited a blue shift as compared to its bulk counterpart, which is an indication of quantum confinement effect. The photoluminescence spectra of CdSe QDs confirmed that the particles are poly-dispersed and possess enhanced luminescent property, depending upon the chemical nature of capping agents. The QDs have been characterized by Fourier-transform infrared spectroscopy, atomic absorption spectroscopy and transmission electron microscopy. Further, antimicrobial activity of as-prepared QDs has also been investigated using the disk diffusion method.     

Fabrication and Characterization of Silk-Fibroin-Coated Quantum Dots

We report a novel technique of directly coating colloidal CdSe/ZnS core/shell quantum dots (QDs) with silk fibroin (SF), a protein derived from the Bombyx mori silk worm. The approach results in protein-modified QDs with little or no particle aggregation, and mitigates the issue of biocompatibility. QDs have desirable optical properties, such as narrow-band emission, broadband absorption, high quantum yield, and high resistance to photobleaching. SF is a fibrous protein polymer with a biomimetic peptide sequence, water and oxygen permeability, low inflammatory response, no thrombogenecity, and cellular biocompatibility, which are desirable properties for in vivo delivery. Combining the unique properties of QDs with the biocompatibility profile of SF, the approach produces particles representing a powerful tool for numerous in vivo and in vitro applications. The design and preparation of these protein-modified QDs conjugates is reported along with functional characterization using luminescence, transmission electron microscope (TEM), and atomic force microscope (AFM). Additionally, we report results obtained using the QDs conjugates as a fluorescent label for bioimaging HEYA8 ovarian cancer cells.      

Energetic alignment in nontoxic SnS quantum dot-sensitized solar cell employing spiro-OMeTAD as the solid-state electrolyte

An environmentally friendly solid-state quantum dot sensitized solar cell (ss-QDSSC) was prepared by combining colloidal SnS QDs as the sensitizer and organic hole scavenger spiro-OMeTAD (2,2′,7,7′-tetrakis-(N,N-di-p-methoxyphenylamine)9,9′-spirobifluorene) as the solid-state electrolyte, and the energy alignment of SnS and TiO₂ was investigated. The bandgap of colloidal SnS QDs increased with decreasing particle size from 14 to 4 nm due to an upshift of the conduction band and a downshift of the valence band. In TiO₂/SnS heterojunctions, the conduction band minimum (CBM) difference between TiO₂ and SnS was as large as ～0.8 eV; this difference decreased with decreasing particle size, but was sufficient for electron injection from SnS nanoparticles of any size into TiO₂. Meanwhile, the sensitizer regeneration driving force, that is, the difference between the valence band maximum (VBM) of SnS and the work function of the electrolyte, showed an opposite behaviour with the SnS size due to a downward shift of the SnS VB. Consequently, smaller SnS QDs should result in a more efficient charge transfer in heterojunctions, revealing the advantages of QDs vs larger particles as sensitizers. This prediction was confirmed by the improved photovoltaic performance of ss-QDSSCs modified with SnS nanoparticles, which peaked for 5–6 nm sized SnS nanoparticles due to the balance between electron injection and sunlight absorption.     

Toxicity of CdTe Quantum Dots on Yeast Saccharomyces Cerevisiae

Along with the widespread development of their bioapplications, concerns about the biosafety of quantum dots (QDs) have increasingly attracted intensive attention. This study examines the toxic effect and subcellular location of cadmium telluride (CdTe) QDs with different sizes against yeast Saccharomyces cerevisiae. The innovative approach is based on the combination of microcalorimetric, spectroscopic, electrochemical, and microscopic methods, which allows analysis of the toxic effect of CdTe QDs on S. cerevisiae and its mechanism. According to the values of the half inhibitory concentration (IC(50) ), CdTe QDs exhibit marked cytotoxicity in yeast cells at concentrations as low as 80.81 nmol L(-1) for green-emitting CdTe QDs and 17.07 nmol L(-1) for orange-emitting CdTe QDs. QD-induced cell death is characterized by cell wall breakage and cytoplasm blebbing. These findings suggest that QDs with sizes ranging from 4.1 to 5.8 nm can be internalized into yeast cells, which then leads to QD-induced cytotoxicity. These studies provide valuable information for the design and development of aqueous QDs for biological applications.