纳米金兔抗猪IgG化学发光探针的制备及表征

采用还原法制备了尺寸大小分别为4、14、30、41 nm的纳米金,优化了不同尺寸的纳米金兔抗猪IgG化学发光探针的制备条件。采用紫外-可见吸收光谱、透射电子显微镜和化学发光的方法对该探针进行了表征。结果表明纳米金兔抗猪IgG化学发光探针具有很好的单分散性和稳定性。本研究为纳米金兔抗猪IgG化学发光探针的应用奠定了基础。     

Determination of gold and palladium in rocks and ores by atomic absorption spectrometry using two-stage probe atomization

The efficiency of two-stage probe atomization for the determination of gold and palladium in geological samples by electrothermal atomic absorption spectrometry is studied. The effects of temperature–time program and the position of the probe in an atomizer on the fractionation of sample components and the magnitude of the analytical signal are studied. It is demonstrated that gold and palladium can be quantitatively determined by atomic absorption spectrometry in rocks and ores, using a two-stage probe atomization with the limits of detection for gold and palladium 0.01 and 0.04 g/t, respectively.     

探针-石墨炉原子吸收光谱法测定岩石矿物中的金

本文采用自制探针系统对探针-石墨炉原子吸收光谱法测金的性能,探针系统的制作,探针的处理,探针原子化法的检测限,灵敏度及抗干扰能力进行了详细的研究,探针原子化的各项指标均优于常规的管壁原子化法。采用本法可不必分离基体物质,直接测定地质样品中的金,所得结果与萃取原子吸收法的结果相吻合。     

Electrochemical detection of human ferritin based on gold nanorod reporter probe and cotton thread immunoassay device

In this study, a natural cotton thread immunoassay device combined with gold nanorod (GNR) reporter probe is developed for the rapid, sensitive and quantitative electrochemical determination of human ferritin, a lung cancer related biomarker. Human ferritin as an analyte and a pair of monoclonal antibodies are used to demonstrate the proof-of-concept on the cotton thread immunoassay device. An enhancement of the sensitivity is achieved by using gold nanorod as an electroactive report probe compared with a traditional gold nanoparticle (GNP) report probe. The device was capable of measuring 1.58 ng/mL ferritin in 30 min by anodic stripping voltammetry (ASV) testing, which meet the requirement for clinical diagnosis.     

Selective decomposition of nucleic acids by laser irradiation on probe-tethered gold nanoparticles in solution

We have developed a new method for selective decomposition of nucleic acids. The method utilizes a high temperature and pressure region (HTP region, hereafter) around a gold nanoparticle, which was generated when the gold nanoparticle was irradiated with a pulsed laser in aqueous solution. A probe DNA molecule whose sequence was complementary to a part of a target DNA molecule was bound to the gold nanoparticle surface. In a solution containing both the target and non-target DNA molecules, the gold nanoparticle selectively attached to the target DNA through hybridization of the probe DNA. When the gold nanoparticle was excited by a pulsed laser, the HTP region was generated in the close vicinity of the gold nanoparticle and then the target DNA molecules inside of this region were decomposed. The non-target DNA molecules having no part complementary to the probe DNA were scarcely decomposed by laser irradiation. When the gold nanoparticle was excited by an intense laser, the non-target DNA molecules were also decomposed, because some of them were located inside the inflated HTP region. We discussed the mechanism of the decomposition of the DNA molecules by the HTP region.     

Dendrimer-based biosensor for chemiluminescent detection of DNA hybridization

We report on a highly sensitive chemiluminescent (CL) biosensor for the sequenc-specific detection of DNA using a novel bio barcode DNA probe modified with gold nanoparticles that were covered with a dendrimer. The modified probe is composed of gold nanoparticles, a dendrimer, the CL reagent, and the DNA. The capture probe DNA was immobilized on magnetic beads covered with gold. It first hybridizes with the target DNA and then with one terminal end of the signal DNA on the barcoded DNA probe. CL was generated by adding H₂O₂ and Co(II) ions as the catalyst. The immobilization of dendrimer onto the gold nanoparticles can significantly enhance sensitivity and gives a detection limit of 6 fmol L^-1 of target DNA.

金电极上L-半胱氨酸短链分子自组装单层的电化学性能和拉曼光谱研究

用循环伏安法分别测定了金电极表面L-半胱氨酸(L-Cys)和十二硫醇自组装单分子层的电化学行为, 实验发现虽然单层结构排列致密, 但并不能有效地阻碍铁氰化钾与电极间异相电子转移过程, 同时观察到十二烷基硫醇自组装层能较好地阻碍电子转移作用. 运用表面增强拉曼散射光谱技术, 以十二烷基硫醇作为缺陷探针, 从分子水平上证实了L-半胱氨酸自组装单层的稳定性和致密性.     

Characterization of single- and double-stranded DNA on gold surfaces

Single- and double-stranded deoxy ribonucleic acid (DNA) molecules attached to self-assembled monolayers (SAMs) on gold surfaces were characterized by a number of optical and electronic spectroscopic techniques. The DNA-modified gold surfaces were prepared through the self-assembly of 6-mercapto-1-hexanol and 5'-C(6)H(12)SH -modified single-stranded DNA (ssDNA). Upon hybridization of the surface-bound probe ssDNA with its complimentary target, formation of double-stranded DNA (dsDNA) on the gold surface is observed and in a competing process, probe ssDNA is desorbed from the gold surface. The competition between hybridization of ssDNA with its complimentary target and ssDNA probe desorption from the gold surface has been investigated in this paper using X-ray photoelectron spectroscopy, chronocoulometry, fluorescence, and polarization modulation-infrared reflection absorption spectroscopy (PM-IRRAS). The formation of dsDNA on the surface was identified by PM-IRRAS by a dsDNA IR signature at approximately 1678 cm(-)(1) that was confirmed by density functional theory calculations of the nucleotides and the nucleotides' base pairs. The presence of dsDNA through the specific DNA hybridization was additionally confirmed by atomic force microscopy through colloidal gold nanoparticle labeling of the target ssDNA. Using these methods, strand loss was observed even for DNA hybridization performed at 25 degrees C for the DNA monolayers studied here consisting of attachment to the gold surfaces by single Au-S bonds. This finding has significant consequence for the application of SAM technology in the detection of oligonucleotide hybridization on gold surfaces.     

QCM DNA biosensor for the diagnosis of a fish pathogenic virus VHSV

Viral haemorrhagic septicaemia (VHS) is one of the most serious viral diseases damaging both fresh and marine fish species. VHS caused by VHSV and diagnosis of VHSV has been dependent on the conventional methods, such as cell culture and RT-PCR, which takes a few days or several hours. This study demonstrates a rapid and sensitive QCM biosensor for diagnosis of VHSV infection in fish. The QCM biosensor was developed to detect a main viral RNA encoding G protein in VHSV using the specific DNA probe. To maximize the sensitivity of the biosensor, we prepared three different DNA probes which modified 3′ end of DNA by thiol, amine, or biotin and compared three different immobilisation methods on quartz surface coated with gold: immobilisation of thiol labelled probe DNA on naked gold surface, immobilisation of amino labelled probe DNA on gold surface prepared as carboxyl chip using MPA followed by EDC/NHS activation, and immobilisation of biotin labelled probe DNA on gold surface after immobilising avidin on carboxyl chip prior to biotin. As a result, immobilisation method using avidin-biotin interaction was most efficient to immobilise probe DNA and to detect target DNA. The QCM biosensor system using biotinylated probe DNA was stable enough to withstand 32 times of repeated regenerations and the detection limit was 0.0016 μM. Diagnosis using the QCM biosensor system was more sensitive and much faster than a conventional RT-PCR analysis in detecting the viral RNA.     

DNA-Templated Gold Nanoclusters for Fluorescence Resonance Energy Transfer-Based Human Serum Albumin Detection

Journal of Analytical Chemistry - A new method for the determination of human serum albumin (HSA) was developed using gold nanoclusters (AuNCs) as a probe. DNA-templated gold nanoclusters with...     

Imaging mass spectrometry of gold nanoparticles in a tissue section as an immunohistochemical staining mass probe

For analysis of low abundance peptides in a tissue section, immunohistochemical staining through antibody‐antigen interaction is a usual technique. The antibody is conjugated with a probe moiety that aids in highly sensitive detection. Gold nanoparticles, which show excellent chemical stability and variation of surface modifications, are expected to act as a sensitive mass probe to desorb gold ions (Au⁺, Au₂⁺, Au₃⁺) that are distinguishable from fragment ions from organic molecules. Here, green fluorescent proteins (GFP) in a tissue section of a transgenic zebrafish were detected by the gold mass probe conjugated with antibodies. Due to the efficient ionization and desorption of gold ions, imaging mass spectrometry of Au₂⁺ ions indicated the distribution of gold nanoparticles stained in a tissue section, and the mass signal distribution was consistent with the area where the GFP‐expressing cells were distributed. Conventional immunofluorescence techniques showed intense autofluorescence that come from intrinsic fluorophores in the tissue section. In contrast, the gold nanoparticles acted as an immunostaining mass probe that displayed significantly lower background signals.     

Probe accessibility effects on the performance of electrochemical biosensors employing DNA monolayers

Surface-confined DNA probes are increasingly used as recognition elements (or presentation scaffolds) for detection of proteins, enzymes, and other macromolecules. Here we demonstrate that the density of the DNA probe monolayer on the gold electrode is a crucial determinant of the final signalling of such devices. We do so using redox modified single-stranded and double-stranded DNA probes attached to the surface of a gold electrode and measuring the rate of digestion in the presence of a non-specific nuclease enzyme. We demonstrate that accessibility of DNA probes for binding to their macromolecular target is, as expected, improved at lower probe densities. However, with double-stranded DNA probes, even at the lowest densities investigated, a significant fraction of the immobilized probe is inaccessible to nuclease digestion. These results stress the importance of the accessibility issue and of probe density effects when DNA-based sensors are used for detection of macromolecular targets.     

Multi-walled Carbon Nanotubes and Gold Nanorod Decorated Biosensor for Detection of microRNA-126

In this article, we introduced a novel electrochemical biosensor for the detection of microRNA-126. The biosensor utilizes a hybridization assay combined with multi-walled carbon nanotubes and gold nanorod-decorated screen-printed carbon electrodes. For electrode preparation, gold nanorods were first immobilized onto the surface of bare and multi-walled carbon nanotube-modified screen-printed carbon electrodes, and the thiol tagged-capture probe was immobilized on the electrode surface through gold and thiol group interaction. After the immobilization, thiol tagged-capture probe hybridized with the target sequence. Under optimum conditions, we determined limit of detection (LOD) and limit of quantification (LOQ) as high as 11 nM and 36 nM, respectively.     

An aptamer-based assay for thrombin via structure switch based on gold nanoparticles and magnetic nanoparticles

An aptamer-based assay for thrombin with high specificity and sensitivity was presented. In the protocol, the aptamer for thrombin was immobilized on magnetic nanoparticle, and its complementary oligonucleotide was labeled with gold nanoparticles, then the aptamer was hybridized with the complementary oligonucleotide to form the duplex structure as a probe, this probe could be used for the specific recognition for thrombin. In the presence of thrombin, the aptamer prefer to form the G-quarter structure with thrombin, resulting in the dissociation of the duplex of the probe and the release of the gold labeled oligonucleotide. Upon this, we were able to detect thrombin through the detection of the electrochemical signal of gold nanoparticles. The strategy combines with the high specificity of aptamer and the excellent characteristics of nanoparticles. This assay is simple, rapid, sensitive and highly specific, it does not require labeling of thrombin, and it could be applied to detect thrombin in complex real sample. The method shows great potential in other protein analysis and in disease diagnosis.     

A bi-ligand co-functionalized gold nanoparticles-based calcium ion probe and its application to the detection of calcium ions in serum

In this study, the use of bi-ligand co-functionalized gold nanoparticles in a highly selective and sensitive colorimetric probe for Ca(2+) ions is demonstrated and this probe also determined the concentrations of Ca(2+) ions in serum samples.     

基于工具酶信号放大技术电化学发光法检测凝血酶

利用巯基乙胺将合成的金纳米粒子氨基化;基于纳米粒子负载羧基化的联吡啶钌和巯基DNA制得电化学发光信号探针;采用酶循环信号放大技术,获得大量含新增DNA的溶液来捕获信号探针;以金电极为载体,将巯基DNA自组装到电极表面,依次杂交互补DNA和信号探针,构建电化学发光生物传感器.在优化的条件下,此传感器对凝血酶具有良好的响应,在3.0× 10-13～6.0×10-11 mol/L范围内,凝血酶的浓度与发光强度呈良好的线性关系,检出限为1.8× 10-13 mol/L(3a).采用酶切循环放大技术制备的生物传感器具有灵敏度高,选择性和重现性良好等特点.     

Sequence-specific detection of femtomolar DNA via a chronocoulometric DNA sensor (CDS): effects of nanoparticle-mediated amplification and nanoscale control of DNA assembly at electrodes

We herein report a novel nanoparticle-based electrochemical DNA detection approach. This DNA sensor is based on a "sandwich" detection strategy, which involves capture probe DNA immobilized on gold electrodes and reporter probe DNA labeled with gold nanoparticles that flank the target DNA sequence. Electrochemical signals are generated by chronocoulometric interrogation of [Ru(NH(3))(6)](3+) that quantitatively binds to surface-confined capture probe DNA via electrostatic interactions. We demonstrated that the incorporation of a gold nanoparticle in this sensor design significantly enhanced the sensitivity and the selectivity. Nanoscale control of the self-assembly process of DNA probes at gold electrodes further increased the sensor performance. As a result of these two combined effects, this DNA sensor could detect as low as femtomolar (zeptomoles) DNA targets and exhibited excellent selectivity against even a single-base mismatch. In addition, this novel DNA sensor showed fairly good reproducibility, stability, and reusability.     

Homogeneous DNA Detection Based on Fluorescence Quenching by Nanoparticles in Single-step Format :Target-Induced Configuration Transform

A new strategy for homogeneous detection of DNA hybridization in single-step format was developed based on fluorescence quenching by gold nanoparticles. The gold nanoparticle is functionalized with 5’-thiolated 48-base oligonucleotide (probe sequence), whose 3’-terminus is labeled with fluorescein (FAM), a negatively charged fluorescence dye. The oligonucleotide adopts an extended configuration due to the electrostatic repulsion between negatively charged gold nanoparticle and the FAM-attached probe sequence. After addition of the complementary target sequence, specific DNA hybridization induces a conformation change of the probe from an extended structure to an arch-like configuration, which brings the fluorophore and the gold nanoparticle in close proximity. The fluorescence is efficiently quenched by gold nanoparticles. The fluorescence quenching efficiency is related to the target concentration, which allows the quantitative detection for target sequence in a sample. A linear detection range from 1.6 to 209.4 nmol/L was obtained under the optimized experimental conditions with a detection limit of 0.1 nmol/L. In the assay system, the gold nanoparticles act as both nanoscaffolds and nanoquenchers. Furthermore, the proposed strategy, in which only two DNA sequences are involved, is not only different from the traditional molecular beacons or reverse molecular beacons but also different from the commonly used sandwich hybridization methods. In addition, the DNA hybridization detection was achieved in homogenous solution in a single-step format, which allows real-time detection and quantification with other advantages such as easy operation and elimination of washing steps.     

Ultrasensitive SERS Detection of Lysozyme by a Target‐Triggering Multiple Cycle Amplification Strategy Based on a Gold Substrate

An ultrasensitive surface enhanced Raman scattering (SERS) method has been designed to selectively and sensitively detect lysozyme. The gold chip as the detection substrate, the aptamer‐based target‐triggering cascade multiple cycle amplification, and gold nanoparticles (AuNPs) bio‐barcode Raman probe enhancement on the gold substrate are employed to enhance the SERS signals. The cascade amplification process consists of the nicking enzyme signaling amplification (NESA), the strand displacement amplification (SDA), and the circular‐hairpin‐assisted exponential amplification reaction (HA‐EXPAR). With the involvement of an aptamer‐based probe, two amplification reaction templates, and a Raman probe, the whole circle amplification process is triggered by the target recognition of lysozyme. The products of the upstream cycle (NESA) could act as the “DNA trigger” of the downstream cycle (SDA and circular HA‐EXPAR) to generate further signal amplification, resulting in the immobility of abundant AuNPs Raman probes on the gold substrate. “Hot spots” are produced between the Raman probe and the gold film, leading to significant SERS enhancement. This detection method exhibits excellent specificity and sensitivity towards lysozyme with a detection limit of 1.0×10^?15 M . Moreover, the practical determination of lysozyme in human serum demonstrates the feasibility of this SERS approach in the analysis of a variety of biological specimens.     

Fabrication of a Sensitive Impedance Biosensor of DNA Hybridization Based on Gold Nanoparticles Modified Gold Electrode

In this work, we report on the preparation of a simple, sensitive DNA impedance sensor. Firstly gold nanoparticles were electrodeposited on the surface of a gold electrode, and then probe DNA was immobilized on the surface of gold nanoparticles through a 5′‐thiol‐linker. Electrochemical impedance spectroscopy (EIS) was used to investigate probe DNA immobilization and hybridization. Compared to the bare gold electrode, the gold nanoparticles modified electrode could improve the density of probe DNA attachment and the sensitivity of DNA sensor greatly. The difference of electron transfer resistance (ΔR_et) was linear with the logarithm of complementary oligonucleotides sequence concentrations in the range of 2.0×10^?12 to 9.0×10^?8 M, and the detection limit was 6.7×10^?13 M. In addition, the DNA sensor showed a fairly good reproducibility and stability during repeated regeneration and hybridization cycles.