Direct electrochemistry of glucose oxidase and electrochemical biosensing of glucose on quantum dots/carbon nanotubes electrodes

Because of their unique chemical, physical and electronic properties, Quantum dots (QDs) and carbon nanotubes (CNTs) are now extremely attractive and important nanomaterials in bioanalytical applications. In this work, CdTe QDs with the size of about 3 nm were prepared and a novel electrochemical biosensing platform of glucose based on CdTe/CNTs electrode was explored. This CdTe/CNTs electrode was prepared by first mixing CdTe QDs, CNTs, Nafion, and glucose oxidase (GOD) in appropriate amounts and then modifying this mixture on the glass carbon electrode (GC). Transmission electron microscopy (TEM) was used to observe the dispersion of CdTe QDs on carbon nanotubes and cyclic voltammetry (CV) was used to investigate the electrochemical behavior of the CdTe/CNTs electrode. A pair of well-defined quasi-reversible redox peaks of glucose oxidase were obtained at the CdTe/CNTs based enzyme electrode by direct electron transfer between the protein and the electrode. The immobilized glucose oxidase could retain bioactivity and catalyze the reduction of dissolved oxygen. Due to the synergy between the CdTe QDs and CNTs, this novel biosensing platform based on QDs/CNTs electrode responded even more sensitively than that based on GC electrode modified by CdTe QDs or CNTs alone. The inexpensive, reliable and sensitive sensing platform based on QDs/CNTs electrode provides wide potential applications in clinical, environmental, and food analysis.     

Spectroscopic investigations on the conformational changes of lysozyme effected by different sizes of N‐acetyl‐l‐cysteine‐capped CdTe quantum dots

The effect of N‐acetyl‐l ‐cysteine‐capped CdTe quantum dots (NAC‐CdTe QDs) with different sizes on lysozyme was investigated by isothermal titration calorimetry (ITC), enzyme activity assays, and multi‐spectroscopic methods. ITC results proved that NAC‐CdTe QDs can spontaneously bind with lysozyme and hydrophobic force plays a major role in stabilizing QDs–lysozyme complex. Multi‐spectroscopic measurements revealed that NAC‐CdTe QDs caused strong quenching of the lysozyme's fluorescence in a size‐dependent quenching manner. Moreover, the changes of secondary structure and microenvironment in lysozyme caused by the NAC‐CdTe QDs were higher with a bigger size. The results of enzyme activity assays showed that the interaction between lysozyme and NAC‐CdTe QDs inhibited the activity of lysozyme and the inhibiting effect was in a size‐dependent manner. Based on these results, we conclude that NAC‐CdTe QDs with larger particle size had a larger impact on the structure and function of lysozyme.     

Interaction and energy transfer studies between bovine serum albumin and CdTe quantum dots conjugates: CdTe QDs as energy acceptor probes

In this paper, a systematic investigation of the interaction of bovine serum albumin (BSA) with water‐soluble CdTe quantum dots (QDs) of two different sizes capped with carboxylic thiols is presented based on steady‐state and time‐resolved fluorescence measurements. Efficient Förster resonance energy transfer (FRET) was observed to occur from BSA donor to CdTe acceptor as noted from reduction in the fluorescence of BSA and enhanced fluorescence from CdTe QDs. FRET parameters such as Förster distance, spectral overlap integral, FRET rate constant and efficiency were determined. The quenching of BSA fluorescence in aqueous solution observed in the presence of CdTe QDs infers that fluorescence resonance energy transfer is primarily responsible for the quenching phenomenon. Bimolecular quenching constant (k_q) determined at different temperatures and the time‐resolved fluorescence data provide additional evidence for this. The binding stoichiometry and various thermodynamic parameters are evaluated by using the van ‘t Hoff equation. The analysis of the results suggests that the interaction between BSA and CdTe QDs is entropy driven and hydrophobic forces play a key role in the interaction. Binding of QDs significantly shortened the fluorescence lifetime of BSA which is one of the hallmarks of FRET. The effect of size of the QDs on the FRET parameters are discussed in the light of FRET parameters obtained.     

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The effect of 3‐mercaptopropionic acid (MPA)‐capped CdTe quantum dots (QDs) on lysozyme was systematically investigated by spectroscopic methods, enzyme activity assay, and calorimetry techniques. Results show that the MPA‐capped CdTe QDs binded to lysozyme through van der Walls forces and hydrogen bondings, causing the decrement of α‐helical content (～7%) and increment of β‐sheet content (～11%) of lysozyme. The binding caused static quenching of the fluorescence, while the microenvironment of aromatic amino acid residues did not show any significant alteration. The lysozyme activity was affected by the increasing exposure of QDs, it was inhibited to 53.77% under a 6 × 10^?7 M exposure compared with the control group. This work will provide direct evidence about enzyme toxicity of QDs to lysozyme in vitro .     

Fluorescence quenching investigation on the interaction of glutathione‐CdTe/CdS quantum dots with sanguinarine and its analytical application

Water‐soluble glutathione (GSH)‐capped core/shell CdTe/CdS quantum dots (QDs) were synthesized. In pH 5.4 sodium phosphate buffer medium, the interaction between GSH‐CdTe/CdS QDs and sanguinarine (SA) was investigated by spectroscopic methods, including fluorescence spectroscopy and ultraviolet‐visible absorption spectroscopy. Addition of SA to GSH‐CdTe/CdS QDs results in fluorescence quenching of GSH‐CdTe/CdS QDs. Quenching intensity was in proportion to the concentration of SA in a certain range. Investigation of the quenching mechanism, proved that the fluorescence quenching of GSH‐CdTe/CdS QDs by SA is a result of electron transfer. Based on the quenching of the fluorescence of GSH‐CdTe/CdS QDs by SA, a novel, simple, rapid and specific method for SA determination was proposed. The detection limit for SA was 3.4 ng/mL and the quantitative determination range was 0.2–40.0 µg/mL with a correlation coefficient of 0.9988. The method has been applied to the determination of SA in synthetic samples and fresh urine samples of healthy human with satisfactory results. Copyright © 2013 John Wiley & Sons, Ltd.     

Photoluminescence properties of hybrid SiO2‐coated CdTe/CdSe quantum dots

Hybrid SiO₂‐coated CdTe/CdSe quantum dots (QDs) were prepared using CdTe/CdSe QDs prepared by hydrothermal synthesis. A CdSe interlayer made CdTe/CdSe cores with unique type II heterostructures. The hybrid SiO₂‐coated CdTe/CdSe QDs revealed excellent photoluminescence (PL) properties compared with hybrid SiO₂‐coated CdTe QDs. Because of the existence of spatial separations of carriers in the type II CdTe/CdSe core/shell QDs, the hybrid QDs had a relatively extended PL lifetime and high stability in phosphate‐buffered saline buffer solutions. This is ascribed to the unique components and stable surface state of hybrid SiO₂‐coated CdTe/CdSe QDs. During the stabilization test in phosphate‐buffered saline buffer solutions, both static and dynamic quenching occurred. The quenching mechanism of the hybrid QDs was not suited with the Stern–Volmer equation. However, the relative stable surface of CdTe/CdSe QDs resulted in lower degradation and relative high PL quantum yields compared with hybrid SiO₂‐coated CdTe QDs. As a result, hybrid SiO₂‐coated CdTe/CdSe QDs can be used in bioapplications. Copyright © 2013 John Wiley & Sons, Ltd.     

Quantum dots as nano plug-in's for efficient NADH resonance energy routing

The routing of fluorescent signals from NADH to quantum dots (QDs) has been a subject of extensive research for FRET based applications. In the present study, the spectral cross talk of NAD(+)/NADH with QDs was used to monitor the reaction of NAD(+)-dependent dehydrogenase enzyme. CdTe QD may undergo dipolar interaction with NADH as a result of broad spectral absorption due to multiple excitonic states resulting from quantum confinement effects. Thus, non-radiative energy transfer can take place from NADH to CdTe QD enhancing QDs fluorescence. Energy routing assay of NADH-QD was applied for detection of formaldehyde as a model analyte in the range 1000-0.01 ng/mL by the proposed technique. We observed proportionate quenching of CdTe QD fluorescence by NAD(+) and enhancement in the presence of NADH formed by various concentrations of enzyme (0.028-0.4 U). Hence, it was possible to detect formaldehyde in the range 1000-0.01 ng/mL with a limit of detection (LOD) at 0.01 ng/mL and regression coefficient R(2)=0.9982. Therefore, a unique optical sensor was developed for the detection of the formaldehyde in sensitive level based on the above mechanism. This method can be used to follow the activity of NAD(+)-dependent enzymes and detection of dehydrogenases in general.     

Molecular spectroscopic studies on the interactions of rhein and emodin with thioglycolic acid‐capped core/shell CdTe/CdS quantum dots and their analytical applications

Water‐soluble thioglycolic acid (TGA)‐capped core/shell CdTe/CdS quantum dots (QDs) were synthesized. The interactions of rhein and emodin with TGA‐CdTe/CdS QDs were evaluated by fluorescence and ultraviolet‐visible absorption spectroscopy. Experimental results showed that the high fluorescence intensity of TGA‐CdTe/CdS QDs could be effectively quenched in the presence of rhein (or emodin) at 570 nm, which may have resulted from an electron transfer process from excited TGA‐CdTe/CdS QDs to rhein (or emodin). The quenching intensity was in proportion to the concentration of both rhein and emodin in a certain range. Under optimized conditions, the linear ranges of TGA‐CdTe/CdS QDs fluorescence intensity versus the concentration of rhein and emodin were 0.09650–60 µg/mL and 0.1175–70 µg/mL with a correlation coefficient of 0.9984 and 0.9965, respectively. The corresponding detection limits (3σ/S) of rhein and emodin were 28.9 and 35.2 ng/mL, respectively. This proposed method was applied to determine rhein and emodin in human urine samples successfully with remarkable advantages such as high sensitivity, short analysis time, low cost and easy operation. Based on this, a simple, rapid and highly sensitive method to determine rhein (or emodin) was proposed. Copyright © 2014 John Wiley & Sons, Ltd.     

CdTe quantum dots (QDs) improve the affinities of baicalein and genistein for human serum albumin in vitro

Baicalein and genistein were studied for the affinities for human serum albumin (HSA) in the presence and absence of three CdTe quantum dots (QDs) with different sizes. Three typical CdTe QDs with maximum emissions of 535 nm (green-emitting, G-QDs), 598 nm (yellow-emitting, Y-QDs), and 654 nm (red-emitting, R-QDs) were tested. The fluorescence intensities of HSA decreased remarkably with increasing concentration of QDs. Baicalein resulted in an obvious blue-shift of the λ_em of HSA from 340 to 334 nm. However, the extents of blue-shifts induced by baicalein and genistein in the presence of QDs were much bigger than that in the absence of QDs. The quenching process of baicalein for HSA was easily affected by the QDs size than that of genistein. QDs increased the quenching constant from 136.97% to 162.24% for baicalein. However, QDs only increased the quenching constants from 20.56% to 32.23% for genistein. G-QDs, Y-QDs, and R-QDs increased the affinities of baicalein for HSA about 3.02%, 6.38% and 9.40%. G-QDs, Y-QDs, and R-QDs increased the affinities of genistein for HSA about 2.56%, 13.46% and 19.44%. The binding affinities of baicalein and genistein for HSA increased with increasing QDs size.     

Aqueous synthesis of CdTe/CdS/ZnS quantum dots and their optical and chemical properties

In this paper, we described a strategy for synthesis of thiol‐coated CdTe/CdS/ZnS (core–shell–shell) quantum dots (QDs) via aqueous synthesis approach. The synthesis conditions were systematically optimized, which included the size of CdTe core, the refluxing time and the number of monolayers and the ligands, and then the chemical and optical properties of the as‐prepared products were investigated. We found that the mercaptopropionic acid (MPA)‐coated CdTe/CdS/ZnS QDs presented highly photoluminescent quantum yields (PL QYs), good photostability and chemical stability, good salt tolerance and pH tolerance and favorable biocompatibility. The characterization of high‐resolution transmission electron microscopy (HRTEM), X‐ray powder diffraction (XRD) and fluorescence correlation spectroscopy (FCS) showed that the CdTe/CdS/ZnS QDs had good monodispersity and crystal structure. The fluorescence life time spectra demonstrated that CdTe/CdS/ZnS QDs had a longer lifetime in contrast to fluorescent dyes and CdTe QDs. Furthermore, the MPA‐stabilized CdTe/CdS/ZnS QDs were applied for the imaging of cells. Compared with current synthesis methods, our synthesis approach was reproducible and simple, and the reaction conditions were mild. More importantly, our method was cost‐effective, and was very suitable for large‐scale synthesis of CdTe/CdS/ZnS QDs for future applications. Copyright © 2010 John Wiley & Sons, Ltd.     

Effects of N‐Acetyl‐L‐Cysteine‐Capped CdTe Quantum Dots on Bovine Serum Albumin and Bovine Hemoglobin: Isothermal Titration Calorimetry and Spectroscopic Investigations

The interactions of N‐acetyl‐L‐cysteine‐capped CdTe quantum dots (QDs) with bovine serum albumin (BSA) and bovine hemoglobin (BHb) were investigated by isothermal titration calorimetry (ITC), fluorescence, synchronous fluorescence, fluorescence lifetime, ultraviolet–visible absorption, and circular dichroism techniques. Fluorescence data of BSA–QDs and BHb–QDs revealed that the quenching was static in every system. While CdTe QDs changed the microenvironment of tryptophan in BHb, the microenvironment of BSA kept unchanged. Adding CdTe QDs affected the skeleton and secondary structure of the protein (BSA and BHb). The ITC results indicated that the interaction between the protein (BSA and BHb) and QDs‐612 was spontaneous and the predominant force was hydrophobic interaction. In addition, the binding constants were determined to be 1.19 × 10⁵ L mol^?1 (BSA–QDs) and 2.19 × 10⁵ L mol^?1 (BHb–QDs) at 298 K. From these results, we conclude that CdTe QDs have a larger impact on the structure of BHb than BSA.     

A novel fluorescent assay for oxytetracycline hydrochloride based on fluorescence quenching of water‐soluble CdTe nanocrystals

A novel assay for oxytetracycline hydrochloride (OTC) based on fluorescence quenching was developed from the interaction between functionalized cadmium telluride quantum dots (CdTe QDs) and OTC. Optimum conditions for the detection of OTC were found after investigating all factors. Under optimum conditions, luminescence of CdTe nanocrystals (λ_ex = 365 nm, λ_em = 562 nm) was quenched by OTC in a concentration‐dependent manner best described by a modified Stern‐Volmer type equation. Good linearity was obtained with a regression coefficient of 0.9999 in the range of 1.34 ~ 13.4 x 10^‐5 mol/L and a limit of detection of 3.08 x 10^‐7 mol/L. In addition, the quenching mechanism was also established. The results imply that the close proximity of OTC‐CdTe was driven by electrostatic attraction and the resulting effective electron transfer from OTC to QDs could be responsible for fluorescence quenching of CdTe‐QDs. Copyright © 2012 John Wiley & Sons, Ltd.     

Luminescence Enhancement of Carboxyl-Coated CdTe Quantum Dots by Silver Nanoparticles

Metallic nanoparticles (NPs) have been used to improve the sensibility of biosensors and bioassays either by enhancing radiative emission or inducing quenching process on fluorescent probes. The aim of this research was to study the interaction of silver and silver-pectin NPs with water-dispersed carboxyl-coated cadmium telluride (CdTe) quantum dots (QDs). Metallic NPs were observed to change the emission of these fluorophores through local field effects. In a solution-base platform, an increase of 82 % was observed for the CdTe emission due to the interaction of QDs and silver-pectin NPs. QDs interaction with silver NPs without pectin was also investigated and a smaller emission enhancement of 20 % was detected. We observed that the NPs’ nature and QDs’ surface charge and concentration are important parameters for NPs-QDs interaction. Moreover, the presence of the pectin polymer shows to be a key component to the observed fluorescence enhancement.     

H2O2‐ and pH‐sensitive CdTe quantum dots as fluorescence probes for the detection of glucose

A novel fluorescence assay system for glucose was developed with thioglycollic acid (TGA)‐capped CdTe quantum dots (QDs) as probes. The luminescence quantum yield of the TGA‐capped CdTe QDs was highly sensitive to H₂O₂ and pH. In the presence of glucose oxidase, glucose is oxidized to yield, gluconic acid and H₂O₂. H₂O₂ and H⁺ (dissociated from gluconic acid) intensively quenched the fluorescence of QDs. The experimental results showed that the quenched fluorescence was proportional to the glucose concentration within the range of 0.01–5.0 mm under optimized experimental conditions. Compared with most of the existing methods, this newly developed system possesses many advantages, including simplicity, low cost, high flexibility, and good sensitivity. Furthermore, no complicated chemical modification of QDs and enzyme immobilization was needed in this system. Copyright © 2012 John Wiley & Sons, Ltd.     

CdTe/ZnS quantum dots as fluorescent probes for ammonium determination

Novel CdTe/ZnS quantum dot (QD) probes based on the quenching effect were proposed for the simple, rapid, and specific determination of ammonium in aqueous solutions. The QDs were modified using 3‐mercaptopropionic acid, and the fluorescence responses of the CdTe/ZnS QD probes to ammonium were detected through regularity quenching. The quenching levels of the CdTe/ZnS QDs and ammonium concentration showed a good linear relationship between 4.0 × 10^?6 and 5.0 × 10^?4 mol/L; the detection limit was 3.0 × 10^?7 mol/L. Ammonium contents in synthetic explosion soil samples were measured to determine the practical applications of the QD probes and a probable quenching mechanism was described. Copyright © 2015 John Wiley & Sons, Ltd.     

Distribution of immobilized enzymes on porous membranes

In this article, the results from a theoretical and experimental investigation of enzyme immobilization in porous membranes are reported. A theoretical model of the immobilization process, which accounts for restricted diffusion of enzyme in the pores of the membrane, has been developed. The model predicts the effect of immobilization kinetics and time of immobilization on the enzyme distribution in the pores of the membrane. The immobilization of glucose oxidase and glucose oxidase-biotin conjugate on porous alumina membranes was experimentally investigated. Enzyme uptake data was correlated to the theory to determine the rate constant of imobilization and the distribution of the enzyme in the pore. Immobilization studies were carried out for enzyme adsorption and for enzyme attachment by covalent coupling. The distribution of enzyme was experimentally studied by assembling five membranes in the diffusion cell. Following immobilization, the membranes were separated and each was assayed for activity. The amount of active enzyme present in each membrane yielded a discrete distribution that compared well with that predicted by theory. (c) 1992 John Wiley & Sons, Inc.     

Biomimetic, mild chemical synthesis of CdTe-GSH quantum dots with improved biocompatibility

Multiple applications of nanotechnology, especially those involving highly fluorescent nanoparticles (NPs) or quantum dots (QDs) have stimulated the research to develop simple, rapid and environmentally friendly protocols for synthesizing NPs exhibiting novel properties and increased biocompatibility. In this study, a simple protocol for the chemical synthesis of glutathione (GSH)-capped CdTe QDs (CdTe-GSH) resembling conditions found in biological systems is described. Using only CdCl(2), K(2)TeO(3) and GSH, highly fluorescent QDs were obtained under pH, temperature, buffer and oxygen conditions that allow microorganisms growth. These CdTe-GSH NPs displayed similar size, chemical composition, absorbance and fluorescence spectra and quantum yields as QDs synthesized using more complicated and expensive methods.CdTe QDs were not freely incorporated into eukaryotic cells thus favoring their biocompatibility and potential applications in biomedicine. In addition, NPs entry was facilitated by lipofectamine, resulting in intracellular fluorescence and a slight increase in cell death by necrosis. Toxicity of the as prepared CdTe QDs was lower than that observed with QDs produced by other chemical methods, probably as consequence of decreased levels of Cd(+2) and higher amounts of GSH. We present here the simplest, fast and economical method for CdTe QDs synthesis described to date. Also, this biomimetic protocol favors NPs biocompatibility and helps to establish the basis for the development of new, "greener" methods to synthesize cadmium-containing QDs.     

碲化镉量子点对PC-12细胞的细胞毒性研究

目的:探讨碲化镉(CdTe)量子点的生物安全性。方法:以水热法制备的、巯基丁二酸(MSA)包被的碲化镉(CdTe)量子点为研究对象,作用于体外培养的鼠神经元细胞(PC-12)。实验设立对照组和不同浓度CdTe QDs作用组。CdTe QDs作用于细胞后,采用倒置荧光显微镜分别观察量子点作用下的细胞形态与细胞对量子点的摄取情况;使用alamarBlue检测CdTe QDs对细胞存活率的影响;Hoechst 33342单染法观察CdTe QDs对PC-12细胞核形态的影响;Annexin V-FITC单染检测CdTe QDs对PC-12细胞凋亡的影响。结果:CdTe量子点作用后,低浓度实验组细胞变化不明显,高浓度实验组中的细胞的形态发生改变,引起细胞肿胀变圆,细胞数量减少,存活率显著下降;在染色下可见明显的细胞凋亡。结论:量子点与细胞接触后会通过胞吞途径进入细胞,汇聚于多种细胞器,对细胞器的功能造成损害,进而引发多种途径的细胞凋亡机制并引发细胞死亡。     

Development of a quantitative relationship between inhibition percentage and both incubation time and inhibitor concentration for inhibition biosensors—theoretical and practical considerations

Theoretical and practical insights into the design and development of immobilised enzyme inhibition biosensors are reported. A general mathematical expression relating the percent of enzyme inhibition (i.e. the analytical signal) to both the inhibitor concentration and the incubation time is presented. The relevant physical, chemical and biochemical parameters required by the model are developed and discussed in terms of the inhibition of acetylcholinesterase by the organophosphorous pesticide, paraoxon. A second enzyme, choline oxidase and an amperometric transducer are used to facilitate the determination acetylcholinesterase inhibitor.     

A highly selective and simple fluorescent sensor for mercury (II) ion detection based on cysteamine‐capped CdTe quantum dots synthesized by the reflux method

Cysteamine (CA)‐capped CdTe quantum dots (QDs) (CA–CdTe QDs) were prepared by the reflux method and utilized as an efficient nano‐sized fluorescent sensor to detect mercury (II) ions (Hg²⁺). Under optimum conditions, the fluorescence quenching effect of CA–CdTe QDs was linear at Hg²⁺ concentrations in the range of 6.0–450 nmol/L. The detection limit was calculated to be 4.0 nmol/L according to the 3σ IUPAC criteria. The influence of 10‐fold Pb²⁺, Cu²⁺ and Ag⁺ on the determination of Hg²⁺ was < 7% (superior to other reports based on crude QDs). Furthermore, the detection sensitivity and selectivity were much improved relative to a sensor based on the CA–CdTe QDs probe, which was prepared using a one‐pot synthetic method. This CA–CdTe QDs sensor system represents a new feasibility to improve the detection performance of a QDs sensor by changing the synthesis method. Copyright © 2014 John Wiley & Sons, Ltd.