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1.
X Cao  M Shen  X Zhang  J Hu  J Wang  X Shi 《Electrophoresis》2012,33(16):2598-2603
PCR has been identified as one of the most important tools in molecular biology and clinical medicine. Improvement of the specificity and efficiency of PCR is often required, and it still remains a great challenge. Here, we introduce the use of dendrimer-entrapped gold nanoparticles (Au DENPs) with different terminal groups prepared using poly(amidoamine) (PAMAM) dendrimers of generation 5 (G5) as a novel class of enhancers to improve the specificity and efficiency of PCR amplification. We show that the optimum concentrations of all the tested Au DENPs are lower than those of the corresponding PAMAM dendrimers without gold nanoparticles (AuNPs). For amine-terminated [(Au(0) )(51.2) -G5.NH(2) ] DENPs, the optimum required concentration is slightly lower than that of G5.NH(2) dendrimers, whereas for glycidol-modified [(Au(0) )(51.2) -G5.NGlyOH] and acetylated [(Au(0) )(51.2) -G5.NHAc] DENPs, the optimum required concentrations are one and three magnitude lower than the corresponding dendrimers, respectively. Our results suggest that the entrapment of AuNPs within the dendrimer interior helps to reserve the 3D spherical morphology of dendrimers, allowing for enhanced interaction with the PCR components. Simultaneously, because of the existence of thermal conductive AuNPs, the enhanced local heat transfer rate may afford decreased chances of mispairing between primers and templates, which is beneficial for enhancing the PCR specificity and efficiency. Therefore, the use of Au DENPs as a novel class of PCR enhancers may enable both improved interaction with the PCR components and the thermal conductivity, which allow them to be used for enhancing different error-prone PCR systems.  相似文献   

2.
This study reports a novel, simple and sensitive immunoassay using fluorescence quenching caused by gold nanoparticles coated with antibody. The method is based on a non-competitive heterogeneous immunoassay of human IgG conducted by the typical procedure of sandwich immunocomplex formation. Goat anti-human IgG was first adsorbed on polystyrene microwells, and human IgG analyte was captured by the primary antibody and then sandwiched by antibody labeled with gold nanoparticles. The sandwich-type immunocomplex was subsequently dissociated by the mixed solution of sodium hydroxide and trisodium citrate, the solution obtained, which contains gold nanoparticles coated with antibody, was used to quench fluorescence. The fluorescence intensity of fluorescein at 517 nm was inversely proportional to the logarithm of the concentration of human IgG in the dynamic range of 10-5000 ng mL−1 with a detection limit of 4.7 ng mL−1. The electrochemical experiments and the UV-vis measurements were applied to demonstrate whether the immunoglod was dissociated completely and whether the gold nanoparticles aggregated after being dissociated, respectively. The proposed system can be extended to detect target molecules such as other kinds of antigen and DNA strands, and has broad potential applications in disease diagnosis.  相似文献   

3.
Xu L  Li B  Jin Y 《Talanta》2011,84(2):558-564
We have demonstrated the design of a new type fluorescence assay based on the inner filter effect (IFE) of gold nanoparticles (AuNPs) on the fluorescence of quantum dots (QDs). With a high extinction coefficient, AuNPs are expected to be capable of functioning as powerful absorbers. QDs with tunable emission wavelength are ideal fluorophores because the emission spectra of the rationally synthesized QDs can perfectly overlap with the absorption band of the absorber. Aminothiols are chosen as the model analytes, and the IFE-based fluorescent method for detection of aminothiols was suggested. Under the optimum conditions, the response is linearly proportional to the concentration of cysteine in the range of 0.05-0.9 μg mL−1. The present IFE-based fluorescent strategy could be also used to detect glutathione and homocysteine. The linear concentration ranges were 0.05-1.0 μg mL−1 for glutathione and 0.01-1.0 μg mL−1 for homocysteine.  相似文献   

4.
<正>The determination method of catechol by fluorescence quenching was developed.The assay was based on the combination of the unique property of gold nanoparticles with tyrosinase enzymatic reaction.In the presence of tyrosinase,the fluorescence of gold nanoparticles was quenched by catechol which can be employed to detect catechol.Under the optimal conditions,a linear range 5.0×10~(-7)-1.0×10~(-3) mol L~(-1) and a detection limit 1.0×10~(-7) mol L~(-1) of catechol were obtained.o-Quinone intermediate produced from the enzymatic catalyzed oxidation of catechol was considered to play the main role in the fluorescence quenching.  相似文献   

5.
A sensitive and convenient strategy was developed for label-free assay of adenosine. The strategy adapted the fluorescence resonance energy transfer property between Rhodamine B doped fluorescent silica nanoparticles (SiNPs) and gold nanoparticles (AuNPs) to generate signal. The different affinities of AuNPs toward the unfolded and folded aptamers were employed for the signal transfer in the system. In the presence of adenosine, the split aptamer fragments react with adenosine to form a structured complex. The folded aptamer cannot be adsorbed on the surface of AuNPs, which induces the aggregation of AuNPs under high ionic concentration conditions, and the aggregation of AuNPs leads to the decrease of the quenching ability. Therefore, the fluorescence intensity of Rhodamine B doped fluorescent SiNPs increased along with the concentration of adenosine. Because of the highly specific recognition ability of the aptamer toward adenosine and the strong quenching ability of AuNPs, the proposed strategy demonstrated good selectivity and high sensitivity for the detection of adenosine. Under the optimum conditions in the experiments, a linear range from 98 nM to 100 μM was obtained with a detection limit of 45 nM. As this strategy is convenient, practical and sensitive, it will provide a promising potential for label-free aptamer-based protein detection.  相似文献   

6.
The high quenching efficiency of metal nanoparticles has facilitated its use as quenchers in molecular beacons. To optimize this system, a good understanding of the many factors that influence molecular beacon performance is required. In this study, molecular beacon performance was evaluated as a function of gold nanoparticle size and its immobilization characteristics. Gold nanoparticles of 4 nm, 15 nm and 87 nm diameter, were immobilized onto glass slides. Each size regime offered distinctive optical properties for fluorescence quenching of molecular dyes that were conjugated to oligonucleotides that were immobilized to the gold nanoparticles. Rigid double stranded DNA was used as a model to place fluorophores at different distances from the gold nanoparticles. The effect of particle size and also the immobilization density of nanoparticles was evaluated. The 4 nm and 87 nm gold nanoparticles offered the highest sensitivity in terms of the change in fluorescence intensity as a function of distance (3-fold improvement for Cy5). The optical properties of the molecular fluorophore was of significance, with Cy5 offering higher contrast ratios than Cy3 due to the red-shifted emission spectrum relative to the plasmon peak. A high density of gold nanoparticles reduced contrast ratios, indicating preference for a monolayer of immobilized nanoparticles when considering analytical performance. Molecular beacon probes were then used in place of the double stranded oligonucleotides. There was a strong dependence of molecular beacon performance on the length of a linker used for attachment to the nanoparticle surface. The optimal optical performance was obtained with 4 nm gold nanoparticles that were immobilized as monolayers of low density (5.7 × 1011 particles cm−2) on glass surfaces. These nanoparticle surfaces offered a 2-fold improvement in analytical performance of the molecular beacons when compared to other nanoparticle sizes investigated. The principles developed in this study would assist in the design of solid phase molecular beacons using gold nanoparticles.  相似文献   

7.
Yu Z  Zhu Y  Zhang Y  Li J  Fang Q  Xi J  Yao B 《Talanta》2011,85(4):1760-1765
In this paper, a nanoliter droplet array based on enzymatic stem-loop probes ligation and SYBR Green real-time PCR for quantification of microRNA was developed. By employing T4 RNA ligase 2 instead of T4 DNA ligase, we designed simplified stem-loop probes to perform microRNA-templated DNA ligation and reduced the non-specific ligation of T4 DNA ligase. SYBR green I dye was employed instead of TaqMan probes in present miniaturized real-time PCR systems. Specifically, we optimized the dosage of SYBR Green I dye in nanoliter droplet and verified the performance of this system by detecting synthetic mir-122 with a 6 logs dynamic range (from 1.5 × 105 to 1.5 × 1010 copies). Linear relationship of the standard curve (R2 = 0.9997) and high PCR amplification efficiency (96.83%) were obtained under the optimized conditions. We detected the expression of mir-122 across five mouse tissues and the result was consistent with that TaqMan microRNA assay. We think this miniaturized real-time PCR platform reduced the detection cost considerably, thus showing the great potential to quantitative biology.  相似文献   

8.
In this study, we used photoluminescence (PL) quenching and transmission electron microscopy (TEM) to study the morphological behavior of hydrogen-bonded (H-bonded) supramolecular assemblies of luminescent H-acceptor polymers and H-donor gold nanoparticles (Au NPs). In fluorescence titration experiments, the lateral Me and MeO substituents on the fluorescent H-acceptor side-chain polymers PBOT1PBOT3 and PBT1PBT3 exhibited different electron-donating capabilities, thereby inducing different degrees of H-bonding and dipole–dipole interactions, as evidenced by effective fluorescence quenching upon the addition of surface-modified Au NPs bearing acid and acid-free surfactants (AuSCOOH and AuSC10, respectively). Among all of our tested nanocomposites, the highest Stern–Volmer quenching constant (KSV) was that obtained from the assembly of AuSCOOH with the homopolymer PBOT1. In addition, we developed fittable exponential equations to predict the values of KSV of other fluorescent polymers (containing various molar ratios of pyridyl conjugated units) when titrated with these NP quenchers. The morphologies observed in the TEM images confirmed that fluorescence quenching resulted from the self-assembly of the supramolecular nanocomposites, mediated by H-bonds between the fluorescent H-acceptors of the polymers and the H-donors of the Au NPs presenting acid-modified surfactants.  相似文献   

9.
In this report we described a highly selective and sensitive iodate sensor. Due to its interaction with fluorescent gold nanoclusters, iodate was capable of oxidizing and etching gold core of the nanoclusters, resulting in fluorescence quenching. Furthermore, it was found that extra iodide ion could enhance this etching process, and even a small amount of iodate could lead to significant quenching. Under an optimized condition, linear relationship between the iodate concentration and the fluorescence quenching was obtained in the range 10 nM–1 μM. The developed iodate sensor was found selective and capable of detecting iodate as low as 2.8 nM. The sensor was then applied for the analysis of iodate in real sample and satisfactory recoveries were obtained.  相似文献   

10.
To evaluate the anti-diabetic effect of Fritillaria cirrhosa gold nanoparticles on Streptozotocin (STZ) stimulated diabetic preclinical models. The albino rats of either sex were equally distributed to five different groups. Group-I represented as Control; Group-II represented as diabetic control (STZ alone); Group-III represented as 10 mg/kg body weight of Fritillaria cirrhosa gold nanoparticles + diabetes; Group-IV represented as 20 mg/kg body weight of Fritillaria cirrhosa gold nanoparticles + diabetes; Group-V represented as 0.1 mg/kg body weight of glibenclamide + diabetes. The animals were killed after the experimental period. The blood and organs samples were gathered and stored for the additional investigations. The gold nanoparticles were inspected via the UV spectrophotometer, HR-TEM, XRD and FT-IR techniques. The bodyweight, kidney and liver weight were tabulated. The food and water consumption were monitored in all the experimental rats. The serum markers, hepatic markers and renal markers were quantified in both normal and investigational rats. The lipid peroxidation and antioxidant status were quantified in the control and experimental rats. The histopathological alterations were also studied in all the experimental animals. The standard drug glibenclamide was used to compare the synthesized AuNPs. The study revealed that the AuNPs treatment restores the serum, hepatic and renal marker in the STZ-challenged diabetic rats. The AuNPs treatment modulates the antioxidants level and decreased the lipid peroxidation by its antioxidant properties. The pathology results revealed that the AuNPs treatment induces the regeneration of islets cells of pancreas in the experimental rats. The research study proved that the Fritillaria cirrhosa AuNPs exerts anti-diabetic properties.  相似文献   

11.
Purpose of the present research work was to evaluate the biological distribution of differently size gold nanoparticles (NP) up on intravenous administration in mice. Another objective was to study effect of particle size on biological distribution of gold NP to enable their diverse applications in nanotechnology. Gold NP of different particle sizes, mainly 15, 50, 100 and 200nm, were synthesized by modifying citrate ion concentration. Synthesized gold nanoparticles were characterized by SEM and their size distribution was studied by particle size analyzer. Gold NP was suspended in sodium alginate solution (0.5%, w/v) and administered to mice (1g/kg, intravenously) [n=3]. After 24h of administration of gold NP, blood was collected under light ether anesthesia, mice were sacrificed by cervical dislocation and various tissues/organs were removed. The tissues were then washed with saline, homogenized and lysed with aqua regia. The determination of gold in samples was carried out quantitatively by inductively coupled plasma mass spectrometry (ICP-MS). SEM study revealed spherical morphology of gold NP with narrow particle size distribution. Biodistribution study revealed gold NPs of all sizes were mainly accumulated in organs like liver, lung and spleen. The accumulation of gold NP in various tissues was found to be depending on particle size. 15nm gold NP revealed higher amount of gold and number of particles in all the tissues including blood, liver, lung, spleen, kidney, brain, heart, stomach. Interestingly, 15 and 50nm gold NP were able to pass blood-brain barrier as evident from gold concentration in brain. Two-hundred nanometers gold NP showed very minute presence in organs including blood, brain, stomach and pancreas. The results revealed that tissue distribution of gold nanoparticles is size-dependent with the smallest 15nm nanoparticles showing the most widespread organ distribution.  相似文献   

12.
In this study, we developed a fluorescence assay for the highly sensitive and selective detection of Hg2+ and Pb2+ ions using a gold nanoparticle (Au NP)-based probe. The Hg–Au and Pb–Au alloys that formed on the Au NP surfaces allowed the Au NPs to exhibit peroxidase-mimicking catalytic activity in the H2O2-mediated oxidation of Amplex UltraRed (AUR). The fluorescence of the AUR oxidation product increased upon increasing the concentration of either Hg2+ or Pb2+ ions. By controlling the pH values of 5 mM tris–acetate buffers at 7.0 and 9.0, this H2O2–AUR–Au NP probe detected Hg2+ and Pb2+ ions, respectively, both with limits of detection (signal-to-noise ratio: 3) of 4.0 nM. The fluorescence intensity of the AUR oxidation product was proportional to the concentrations of Hg2+ and Pb2+ ions over ranges 0.05–1 μM (R2 = 0.993) and 0.05–5 μM (R2 = 0.996), respectively. The H2O2–AUR–Au NP probe was highly selective for Hg2+ (>100-fold) and Pb2+ (>300-fold) ions in the presence of other tested metal ions. We validated the practicality of this simple, selective, and sensitive H2O2–AUR–Au NP probe through determination of the concentrations of Hg2+ and Pb2+ ions in a lake water sample and of Pb2+ ions in a blood sample. To the best of our knowledge, this system is the first example of Au NPs being used as enzyme-mimics for the fluorescence detection of Hg2+ and Pb2+ ions.  相似文献   

13.
We present a novel approach to improve the analytical figures of merit of solid-phase extraction high-performance liquid chromatography (SPE-HPLC) for the analysis of monohydroxy-polycyclic aromatic hydrocarbons in urine samples. The novel alternative substitutes the evaporation step that is currently used in SPE-HPLC methodology with a pre-concentration procedure that extracts metabolites with gold nanoparticles. The analytical potential of the new approach is evaluated with the following six metabolites: 9-hydroxyphenanthrene, 2-hydroxyfluorene, 1-hydroxypyrene, 6-hydroxychrysene, 3-hydroxybenzo[a]pyrene and 4-hydroxybenzo[a]pyrene. We demonstrate that the substitution of the evaporation step with the gold nanoparticles procedure improves the overall recoveries, the relative standard deviations of the average recoveries and the limits of detection of SPE-HPLC analysis. The overall recoveries of the studied metabolites varied from 59.7 ± 3.6% (2-hydroxyfluorene) to 92.3 ± 2.5% (6-hydroxychrysene). The relative standard deviations of the average recoveries were lower than 6%. The limits of detection were at the parts-per-trillion levels and varied from ∼2 pg mL−1 (6-hydroxychrysene) to ∼18 pg mL−1 (2-hydroxyfluorene).  相似文献   

14.
In this work, an active nano-catalyst with gold nanoparticles loaded in hollow mesoporous silica nanospheres (HMSNs/Au) was prepared by a one-pot sol-gel method, in which gold ions were loaded in hollow mesoporous silica spheres followed by sodium alginate reduction. The characterization of the HMSNs/Au were determined by scanning electron microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD), N2 adsorption–desorption isotherms (BET). The high catalytic activity of HMSNs/Au, denoted as apparent turn-over frequency (TOF), was detected by UV-Vis spectrophotometer for the catalytic reduction of 4-nitrophenol (74.5 h?1) and 2-nitrophenol (108.7 h?1) in the presence of sodium borohydride solution due to the small gold nanoparticles size and overall exposure of active sites. It is expected that this ecofriendly approach to prepare inorganic composited nanoparticles as high active catalysts based on hollow mesoporous materials was a promising platform for loading noble metal nanoparticles.  相似文献   

15.
Sui B  Shen L  Jin W 《Talanta》2011,85(3):1609-1613
An ultrasensitive solid-phase fluorescence resonance energy quenching (FREQ) method for determination of 1,4-dihydroxybenzene (DHB) using mercaptosuccinic acid (MSA)-capped CdTe quantum dots (QDs) immobilized on silica nanoparticles (NPs) as donors was developed. In the method, silica NPs were first modified with 3-aminopropyltriethoxysilane (APTS). Then, MSA-capped CdTe QDs were immobilized on the surface of the APTS-modified silica NPs. Finally, DHB in the solution was attached to the empty sites on the surface of silica NPs with QDs through electrostatic interaction. The fluorescence emission of the QDs was quenched by the proximal DHB molecules on the silica NPs. The quenching efficiency of the solid-phase FREQ method was 200-times higher than that of the solution-phase FREQ method. Using the ultrasensitive solid-phase FREQ method, DHB as low as 2.4 × 10−12 mol/L could be detected. The method was applied to quantify trace DHB in water samples.  相似文献   

16.
A facile green biosynthesis method has been successfully developed to prepare gold nanoparticles (AuNPs) of various core sizes (25 ± 7 nm) using a natural biomaterial, eggshell membrane (ESM) at ambient conditions. In situ synthesis of AuNPs-immobilized ESM is conducted in a simple manner by immersing ESM in a pH 6.0 aqueous solution of HAuCl4 without adding any reductant. The formation of AuNPs on ESM protein fibers is attributed to the reduction of Au(III) ions to Au(0) by the aldehyde moieties of the natural ESM fibers. Energy dispersive X-ray spectroscopy, scanning electron microscopy, X-ray photoelectron spectroscopy, and X-ray powder diffraction unambiguously identify the presence of AuNPs on ESM. The effect of pH on the in situ synthesis of AuNPs on ESM has been investigated in detail. The pH of the gold precursor (HAuCl4) solution can influence the formation rate, dispersion and size of AuNPs on ESM. At pH ≤3.0 and ≥7.0, no AuNPs are observed on ESM while small AuNPs are homogeneously dispersed on ESM at pH 4.0-6.0. The optimal pH for AuNPs formation on ESM is 6.0. AuNPs/ESMs are used to immobilize glucose oxidase (GOx) for glucose biosensing. AuNPs on ESM can increase the enzyme activity of GOx. The linear response range of the glucose biosensor is 20 μM to 0.80 mM glucose with a detection limit of 17 μM (S/N = 3). The biosensor has been successfully applied to determine the glucose content in commercial glucose injections. Our work provides a very simple, non-toxic, convenient, and green route to synthesize AuNPs on ESM which is potentially useful in the biosensing field.  相似文献   

17.
In this paper, we demonstrate a simple and sensitive colorimetric detection of cysteine based on the cysteine-mediated color change of ssDNA-stabilized gold nanoparticles (AuNPs). Cysteine is capable of absorbing onto AuNPs surfaces via the strong interaction between its thiol group and gold. ssDNA molecules which stabilize AuNPs against salt-induced aggregation are removed away by cysteine encapsulation on the AuNPs surfaces, resulting in a characteristic color change of AuNPs from red to blue as soon as salt is added. The ratio of absorptions at 640 to 525 nm (A 640/A 525) is linear dependent on the cysteine concentration in the range from 0.1 to 5 μM. Furthermore, amino acids other than cysteine cannot mediate the color change under the identical conditions due to the absence of thiol groups, suggesting the selectivity of the proposed method toward cysteine. The employment of complicated protocols and sophisticated processes such as the preparation of modified AuNPs are successfully avoided in design to realize the simple and low-cost cysteine detection; and the high sensitivity and low cost of the method is favorable for practical applications. Figure In the presence of cysteine, cysteine binds to the AuNPs surface via Au-S bond, spontaneously driving ssDNA molecules away from the nanoparticles, which leads to the AuNPs aggregation under the condition of NaCl introduction, and the corresponding color change from red to blue. However, the presence of other amino acids results in no color change due to the absence of thiol groups. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

18.
Peanut allergic reactions can result from the ingestion of even very small quantities of peanut and represent a severe threat to the health of sensitised individuals. The detection of peanut traces in food products is therefore of prime importance. Peanut traces which can be (unintentionally) present in food products have usually undergone one or more processing steps like roasting and baking. Therefore, methods designed to detect such traces have to be capable of detecting heat-treated peanuts. Commonly used methodologies designed to detect peanut traces in food products are enzyme-linked immunosorbent assays (ELISAs) that detect peanut-specific proteins, and polymerase-chain-reaction (PCR)-based methods targeting peanut-specific DNA. A comparative analysis of such methods was performed and the impact of heat treatment on peanut kernels as well as the impact on a peanut-containing food matrix are investigated. Our results show that heat treatments have a detrimental effect on the detection of peanut with either type of method and that both types of methods are affected in a similar manner.  相似文献   

19.
Synthetic oligopeptides with a tryptophan residue at the C-terminus have been used for the synthesis of gold and silver nanoparticles at pH 11. The tryptophan residue in the peptides is responsible for the reduction of metal ions to the respective metals, possibly through electron transfer. A mechanistic pathway has been proposed to explain the reductive properties of the tryptophan moiety of the peptide based on some spectroscopic techniques, such as UV-visible and fluorescence spectroscopy. This study reveals that some of the peptide molecules are converted to its corresponding ditryptophan, kynurenine form and some cross-linked products, all of which are highly fluorescent species. The resultant peptide-functionalized metal nanoparticles have also been characterized by UV-visible spectroscopy, transmission electron microscopy, and Fourier transform IR spectroscopy and thermogravimatric analysis.  相似文献   

20.
We report our findings that natural flavonoids such as quercetin, daizeol and puerarin can act as reductants for the enlargement of gold nanoparticles (Au-NPs). Consequently, the UV–vis spectra of a solution containing Au-NPs will be gradually changed, and the molecules of the natural herbs can be detected by making use of changes in the UV–visible spectra. Furthermore, we have prepared a self-assembled monolayer modified electrode by modifying cysteamine on a gold substrate electrode, which is further modified by some Au-NP seeds. When the modified electrode is immersed in a solution containing flavonoids and tetrachloroauric acid as a gold source for the growth of the Au-NP seeds, with the increase of the concentration of flavonoids, the Au-NP seeds on the surface of the modified electrode can be enlarged to varying degrees. As a result, the peak currents in the corresponding cyclic voltammograms are inversely decreased, and simultaneously the peak separation is increased. Therefore, an electrochemical method to detect flavonoids is also proposed. Compared with the optical detection method, the electrochemical method has an extraordinarily lower detection limit and a significantly extended detection range. Moreover, the optical and electrochemical experimental results can be also used to assay and compare the relative antioxidant activities of the flavonoids. Figure Enlargement of Au nanoparticles by flavonoids at cysteamine modified electrode  相似文献   

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