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31.
o-Dianisidine (ODA)-H2O2-horseradish peroxidase (HRP) voltammetric enzyme-linked immunoassay system has firstly been used for the detection of tobacco mosaic virus (TMV). HRP catalyzes strongly the oxidation reaction of ODA by H2O2, the product of which produces a sensitive second order derivative linear sweep voltammetric peak at potential of −0.56 V (versus SCE) in Britton–Robinson (BR) buffer. HRP activity has been measured with this voltammetric peak and TMV detected through immunoreaction. The detection limit for HRP is 9.25×10-7 mU l−1 and the linear range is 2.5×10−6–5.0×10−4 mU l−1. The detection limit for the clarified TMV is 0.25 ng ml−1 and the highest dilution ratio detected for the infected leaf sap is 1:8×105. The sensitivity for TMV detection with this method is higher than that with the enzyme-linked immunosorbent spectrophotometric assay (ELISA) using ODA-H2O2-HRP system. The processes of the enzyme-catalyzed reaction and the electro-reduction of the product of the enzyme-catalyzed reaction have been described. 相似文献
32.
为将生物体内微观的蛋白行为可视化并以宏观信号呈现出来对蛋白进行实时、动态分析,借助SNAP-tag蛋白标签技术与有机小分子荧光染料,构建了一系列用于活细胞内实时监测目标蛋白的免洗荧光探针。标签蛋白SNAP-tag能够特异性识别探针中的苄基鸟嘌呤,从而使目标蛋白共价连接上荧光团(萘酰亚胺),携带上荧光信使。此外,由于萘酰亚胺从水环境中被牵引至SNAP-tag蛋白的疏水空腔,其荧光信号呈现出2~13倍的增强。通过SNAP-tag标签蛋白与目标蛋白的融合,该荧光探针实现了对活细胞内线粒体蛋白CoX8A及核内蛋白H2B特异性识别,在免洗条件下完成了对目标蛋白的实时追踪及原位分析。 相似文献
33.
Nitric oxide (NO) is an important mediator responsible for numerous physiological phenomena. Transient levels of NO in biological systems usually range from nanomolar to micromolar concentrations, with a rapid return to basal levels normally seen following these increases. Because NO can diffuse only over a local area in limited time due to such low levels of production and due to its short life-time prior to degradation, high spatial and temporal resolutions are required for direct and continuous NO measurement if the physiological role of NO is to be investigated in any system. For such purposes, analytical methods based on bio-imaging and electrochemical techniques for the measurement of NO are useful. In this paper, we describe the successful application of these methods to a number of biological systems. Specifically, complementary application of these methods demonstrate that it is possible to detect real-time NO production from nervous tissue with high spatial and temporal resolutions. 相似文献
34.
Miyuki Narita Fumio Hamada Mayumi Sato Iwao Suzuki Tetsuo Osa 《Journal of inclusion phenomena and macrocyclic chemistry》1999,34(4):421-430
Flexible hosts, 6A,6B-; 6A,6C-; 6A,6D-; and 6A,6E-bis dansylglycine-modified -cyclodextrins (-1, -2, -3, and -4, respectively) have been synthesized as a sensing molecule for organic guests including terpenoids and bile acids. These host compounds show a pure monomer fluorescence whose intensity is decreased or enhanced upon addition of guest species. The value I/I0, where I and I0 are fluorescence intensities in the presence and absence of a guest and I is I0- I, was used as a parameter of sensitivity. These hosts exhibit highly sensitive and selective molecular recognition ability, particularly, for lithochoic acid, chenodeoxycholic acid, and ursodeoxycholic acid. The behaviors of the appended moieties of these hosts when host–guest complexation occurs are studied by induced circular dichroism (ICD) spectra and fluorescence spectral change on accommodation of a guest. The ICD pattern of these hosts alone or on accommodation of a guest is very similar, indicating that the behavior of the appended moieties are very similar. The guest-induced variations in the fluorescence or ICD intensity suggest that the appended moieties act as a hydrophobic cap that enables the cyclodextrin to form 1 : 1 host–guest complexes. 相似文献
35.
Multichannel homogeneous immunoassay for detection of 2,4,6-trinitrotoluene (TNT) using a microfabricated capillary array electrophoresis chip 总被引:6,自引:0,他引:6
A high-throughput homogeneous immunoassay for the sensitive detection of 2,4,6-trinitrotoluene (TNT) has been developed using radial capillary array electrophoresis microdevices. Samples consisting of equilibrium mixtures of anti-TNT antibody (Ab), fluorescein-labeled TNT, and various concentrations of unlabeled TNT were electrokinetically injected into 48 channels of a radial capillary array electrophoresis microchannel plate. The rapid electrophoretic separation allows us to analyze the equilibrium ratio formed by the competition between the labeled and the unlabeled TNT for Ab binding. The simultaneous parallel TNT separations facilitate determination of a calibration curve for the TNT assay, which has high sensitivity (LOD, 1 ng/mL) and a wide dynamic range (1-300 ng/mL). 相似文献
36.
Summary The retention, selectivity and elution order of fluorescent 1,N6-etheno derivatives of diadenosine polyphosphates and their enzymatic degradation products on octadecyl and phenyl-bonded
silica columns have been studied as a function of mobile phase pH, ionic strength and organic modifier content. Good separations
of the compounds of interest were achieved using mobile phases of around 0.1M potassium phosphate buffers at neutral pH containing
approximately 10% methanol or 4% acetonitrile for C18 columns and 5% methanol or 1.5% acetonitrile for phenyl columns. The data obtained were used to establish isocratic assays
for diadenosine polyphosphate cleaving activities from chromaffin cells using Di(1,N6-ethenoadenosine) polyphosphates as fluorogenic substrate analogues followed by fluorescence detection. 相似文献
37.
38.
Matthew R OatesWilliam Clarke Alden Zimlich IIDavid S Hage 《Analytica chimica acta》2002,470(1):37-50
Various practical and theoretical considerations were examined in the creation and optimization of a high-performance liquid chromatography (HPLC)-based one-site immunometric assay. This method used an HPLC analyte analog column and post-column chemiluminescence detection. The specific analyte chosen as the model for this study was l-thyroxine (also known as T4). In this technique, a sample containing thyroxine was first combined with an excess of anti-T4 antibody Fab fragments that had earlier been conjugated with chemiluminescent acridinium ester labels. After incubation, the mixture was injected onto a column that contained immobilized T4. The amount of thyroxine in the original sample was then determined by measuring the labeled Fab fragments that appeared in the non-retained fraction, or the decrease in excess Fab fragments that were bound to and later eluted from the column. Items considered in creating this assay included the preparation of acridinium ester-labeled Fab fragments, the detection of these fragments with a post-column reactor, and the creation of a suitable immobilized analog column for capturing excess labeled Fab fragments. The final method could measure T4 in standards at clinically-relevant concentrations and provided a response within 1.5 min of sample injection, following a 20-45 min incubation with the labeled Fab fragments. Possible applications of this method include its use in clinical chemistry and the screening of proteomic or combinatorial libraries. 相似文献
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