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ABSTRACT: Here, a double-strand (ds) DNA microarray-based fluorescence assay has been deployed for studying endonuclease functionality and inhibition. The dsDNA microarrays are fabricated by hybridization of the Cy5-labeled oligonucleotides with the immobilized complementary oligonucleotide probes on the glass slide. The microarray displays significant fluorescence decrease in response to endonuclease, which is able to cleave the oligonucleotide moiety of dsDNA. Two endonucleases, EcoRI and BamHI, and four potential inhibitors, doxorubicin hydrochloride (DOX), 5-fluorouracil (5-FU), ethidium bromide (EB) and actinomycin D (ACTD), were selected to address the feasibility of this assay. Enzyme activities of the endonucleases are detected with high specificity down to the limits of 1.1 U mL(-1) for EcoRI and 2.0 U mL(-1) for BamHI, respectively. In addition, BamHI and EcoRI inhibition by the inhibitors are also shown, demonstrating the potential for high-throughput screening for inhibitors.
The Analyst 01/2013; · 4.23 Impact Factor
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ABSTRACT: A simple, sensitive gold nanoparticle (GNP)-based dot-blot immunoassay has been developed for detecting Alzheimer's disease related β-amyloid peptide 1-42 (Aβ(1-42)) down to a 50 pg mL(-1) level in aqueous solution. Practical samples (cerebrospinal fluid (CSF), cell culturing mediums and cell lysates) have been used to demonstrate the ability of the assay.
Chemical Communications 07/2012; 48(67):8392-4. · 6.17 Impact Factor
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ABSTRACT: Herein, we report a phenylboronic acid functionalized gold nanoparticle (GNP)-based colorimetric assay for rapid detection of Staphylococcus aureus (S. aureus) with high sensitivity. In this approach, GNPs can bind to S. aureus by the reaction of phenylboronic acid with the cis-diol configuration in glycans on the bacterial surface, providing a colorimetric readout of the binding event. Using this strategy, we have been able to quantify S. aureus at a concentration of 50 cells per mL (three times the standard deviation divided by the slope of the working curve) in aqueous solution.
Nanoscale 12/2011; 4(2):451-4. · 5.91 Impact Factor
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ABSTRACT: In the present work, a kind of peptide functionalized gold nanoparticle (AuNP) has been synthesized and employed for colorimetric detection of Pb(2+) in both aqueous solution and living cell. The AuNPs are capped by two peptide ligands: glutathione (GSH) and pentapeptide (CALNN). The GSH is used as a functional group for selectively sensing Pb(2+) by coordination reaction, and CALNN is employed as a stabilize ligand for improving the stability of AuNPs under physiological condition, respectively. The AuNP enables to strongly interact with Pb(2+) that leads to distinct color change of solution. Under the optimized molar ratio of GSH to CALNN on the AuNP surface, the colorimetric assay for detecting Pb(2+) in living cell downs to 2.9 fmol Pb(2+)per cell (3 times of standard deviation, 3σ) with linear relationship from 2.9 to 37.7 fmol Pb(2+) per cell. In addition, the method also shows highly selective detection toward Pb(2+) against other common metal ions in both aqueous solution and living cell.
Biosensors & bioelectronics 11/2011; 31(1):505-9. · 5.43 Impact Factor
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ABSTRACT: A novel biosensing strategy has been developed for sensitive detection of β-amyloid 1-42 (Aβ(1-42)) down to the ng mL(-1) level in both aqueous solution and diluted human plasma, which is based on measuring changes in resonance light scattering of streptavidin functionalized gold nanoparticles as a function of the concentration of Aβ(1-42) with a conventional spectrofluorometer.
Chemical Communications 09/2011; 47(33):9339-41. · 6.17 Impact Factor
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ABSTRACT: Here, a three-dimensional (3D) carbohydrate microarray-based plasmon resonance light scattering (RLS) assay has been established for studying carbohydrate-lectin binding with high selectivity. The 3D carbohydrate microarray is fabricated by immobilizing amino-modified carbohydrates on the home-made fourth-generation (G4) NH(2)-terminated poly(amidoamine) dendrimers (PAMAM)-modified substrate. After marking the carbohydrate-lectin binding events by 13 nm peptide-stabilized gold nanoparticles through the biotin-avidin reaction, the 3D microarray can be directly detected by the RLS scanner without the conventional silver enhancement step. The well defined recognition systems: three monosaccharides (Man-α, Glc-α and Gal-β) with two lectins (Con A and RCA 120), have been chosen here to establish the RLS assay, respectively. Quantitative determination of the surface dissociation constants (K(D,surf)) for surface carbohydrates and lectins has been achieved. In addition, inhibition values (i.e. the inhibition constants (K(i)) and the concentrations of inhibitors required to produce 50% inhibition (IC(50))) for inhibitors in solution are also demonstrated by the saccharide competing assays.
The Analyst 09/2011; 136(20):4301-7. · 4.23 Impact Factor
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ABSTRACT: In the post-genomic era, glycomics (the functional study of carbohydrates in living organisms) has come into the forefront of biological research because the interactions of glycoconjugates with proteins not only occur widely in biological processes of cells but also initiate infection of host cells by bacteria and viruses. Microarrays have been reportedly successful in carbohydrate-protein interaction as well as cellular surface glycan profiling. This review provides an overview of recent progress in the development of microarray-based techniques for glycomic studies. The fabrication, application and challenge/bottleneck of glycan/lectin microarrays have been summarized and discussed.
Combinatorial chemistry & high throughput screening 07/2011; 15(1):90-9. · 2.46 Impact Factor
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ChemPhysChem 04/2011; 12(5):909-12. · 3.41 Impact Factor
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ABSTRACT: To develop a novel high-throughput tool for monitoring specific affinity of microbes with lectins, a kind of lectin microarray has been fabricated by immobilizing lectins on epoxide-derivatized glass slides and used to capture microbes. The capturing events are marked by attachment of lectin-conjugated gold nanoparticles followed by silver deposition to enhance the resonance light scattering (RLS) of the particles. The interactions of 16 lectins with four bacteria and one fungus were profiled by this approach. We demonstrated that the gold-nanoparticle-labeled array was suitable for identifying the binding affinity of lectin with bacterium, as well as determining the bacterium with high sensitivity. More importantly, we found that the growth of microbial strains in different culture media resulted in significant changes in their binding affinities with lectins, which might be important to the pathogenesis of the organisms.
Analytical Chemistry 10/2010; 82(22):9240-7. · 5.86 Impact Factor
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ABSTRACT: In present work, a rhodamine 6G (Rh 6G)-incorporated β-cyclodextrin functionalized gold nanoparticle (Rh 6G-CD-AuNP) based fluorescent assay has been successfully developed for recognizing/detecting the structural isomers, α-naphthol and β-naphthol, in aqueous solution. The β-cyclodextrin functionalized gold nanoparticles (CD-AuNPs) are achieved by conjugating the thiolated β-cyclodextrin (SH-β-CD) with AuNPs via S-Au covalent bonds. Rhodamine 6G (Rh 6G) is chosen as a fluorescent probe in this approach because it can be strongly absorbed on the surface of AuNP by noncovalent interaction. After binding with β-CD cavity, the naphthols enable to act as electron transfer quenchers of Rh 6G, which lead to significant fluorescence quenching of the dye. Because of different association ability of naphthol isomers with the β-CD cavity, the assay can selectively distinguish α-naphthol and β-naphthol with reasonable sensitivity. Detection of naphthols down to 8 nM with a dynamic range of nearly three orders of magnitude (0.01-8 μM) for α-naphthol and 50 nM with two orders of magnitude (0.1-20 μM) for β-naphthol is demonstrated, respectively. The ability of the method for detecting the content of α-naphthol or β-naphthol in the different naphthol mixtures has also been evaluated.
Biosensors & bioelectronics 10/2010; 26(5):2329-33. · 5.43 Impact Factor
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ABSTRACT: In this paper, we propose an effective protocol for bacterial detection using antibody microarray-based strategies with gold nanoparticle probes. In this approach, bacteria are captured by immobilized IgGs on the microarray spots. The capturing events are marked by attachment of lectin (e.g. Ricinus Communis Agglutini (RCA) and Concanavalin A (ConA)) conjugated gold nanoparticles followed by silver deposition for signal enhancement. The detection principle is resonance light scattering (RLS). Using this technique, highly selective discrimination of bacterial strain types (i.e., gram-negative, gram-positive and fungus) down to 10(3) cells per assay with a dynamic range of nearly three orders of magnitude are demonstrated. In addition, the capturing process can be inhibited by lipopolysaccharides (LPS), which gives further evidence of the binding mechanism. Furthermore, antibacterial assay with antibacterial drugs (amoxicillin and vancomycin) demonstrates that the approach enables to detect antibacterial efficiency as well as derive MIC (minimum inhibitory concentration) plots.
Talanta 06/2010; 81(4-5):1816-20. · 3.79 Impact Factor
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ABSTRACT: In this paper, a kind of beta-amyloid peptide (Abeta1-16) conjugated gold nanoparticles (Abeta1-16@GNPs) are prepared and employed as colorimetric indicator for studying the interaction of beta-amyloid peptide with metallic ions (e.g. Zn(2+) and Ca(2+)). In the presence of Zn(2+), mono-dispersing Abeta1-16@GNPs enable to form aggregates or attach on the SHG-44 (human glioma cell) cellular surface which results in significant color change of the solution. The experimental results indicate that Zn(2+) can interact with Abeta1-16 and form Zn(2+)-beta-amyloid peptide complexes. In particular, in the presence of Zn(2+), a time-dependent interaction of cells with Abeta1-16@GNPs has been observed that may suggest different expression levels of beta-amyloid peptide related proteins in various cell cycles. In addition, the aggregating/binding process can be easily reversed by adding EDTA, a good chelated ligand of Zn(2+), which gives further proof of the interaction mechanism.
Talanta 03/2010; 80(5):1626-31. · 3.79 Impact Factor
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ABSTRACT: In this technical note, a microarray-based spectroscopic assay with two readout principles, fluorescence and resonance light scattering (RLS), for screening kinase inhibitors has been reported. In this assay, the phosphorylation and inhibition events are marked by biotinylated antiphosphoserinen/antiphosphotyrosine antibodies, and gold nanoparticles are attached to the antibodies by standard avidin-biotin chemistry followed by silver deposition for RLS signal enhancement. The avidin conjugated fluorescein is used as a fluorescent probe. Assays for both serine kinase, the alpha-catalytic subunit of cyclic adenosine 5'-monophosphate (cAMP) dependent protein kinase (PKA), and tyrosine kinase, leukocyte-specific protein tyrosine kinase (LCK), have been developed. The utility of this assay to high-throughput screening was demonstrated with a commercial inhibitor library, a collection of 80 kinase inhibitors, and satisfactory results were obtained. In addition, quantitative determination of binding strength and the inhibiting type (type I) of these inhibitors are also demonstrated by the adenosine 5'-triphosphate (ATP) competing assays.
Analytical Chemistry 03/2010; 82(7):3067-72. · 5.86 Impact Factor
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ABSTRACT: A colorimetric assay based on pentapeptide (CALNN) functionalized gold nanoparticles exhibits high sensitivity and selectivity for detection of aluminium cation (Al(3+)) both in aqueous solution and on living cellular surfaces under physiological condition.
Chemical Communications 02/2010; 46(6):988-90. · 6.17 Impact Factor
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ABSTRACT: Here, we describe the synthesis of peptide- and/or protein-functionalized Fe(2)O(3) core-Au shell (Fe(2)O(3)@Au) nanoparticles for imaging and targeting of living cells. When functionalized with the transmembrane peptide RRRRRRRR (R(8)), the Fe(2)O(3)@Au nanoparticles (R(8)-Fe(2)O(3)@Au) are able to serve as cellular trafficking agents with excellent biocompatibility. The internalization mechanism and delivery efficiency of the R(8)-Fe(2)O(3)@Au nanoparticles have been characterized with dark-field microscopy and fluorescence confocal scanning laser microcopy. Experimental result suggests that the R(8)-Fe(2)O(3)@Au nanoparticles are internalized initially by binding with the membrane-associated proteoglycans on cell surfaces, especially heparan sulfate proteoglycans (HSPGs), following an energy-dependent endocytosis process to enter into living cells. After conjugation with the epidermal growth factor receptor antibody (anti-EGFR), these nanoparticles can also be used for the recognition of cell membrane antigens to specifically label tumor cells.
Nanoscale 02/2010; 2(2):269-76. · 5.91 Impact Factor
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ABSTRACT: To develop a novel high-throughput tool for monitoring carbohydrate-protein interactions, carbohydrate or glycoprotein microarrays have been prepared for binding with lectins. The interaction events are marked by attachment of fluorescent dyes and gold nanoparticles. The attachment of the fluorescent dyes and gold nanoparticles is achieved by standard avidin-biotin chemistry. The detection principle is fluorescence or resonance light scattering (RLS). The electroless deposition of silver onto the gold particles has been employed for RLS signal enhancement. Well-defined recognition systems, three monosaccharides (Man-alpha, Glc-beta, and Gal-beta) or three glycoproteins (Asf, RNase A, and RNase B) with two lectins (ConA and RCA120), are chosen here to establish the microarray-based assay, respectively. Highly selective recognition of carbohydrate-protein down to 25.6 pg/mL for RCA120 in solution and 8 microM for Gal-beta and 32 ng/mL for Asf on the microarray spots is demonstrated.
Methods in molecular biology (Clifton, N.J.) 01/2010; 600:145-53.
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ABSTRACT: To develop a novel high-throughput tool for monitoring carbohydrate–protein interactions, carbohydrate or glycoprotein microarrays
have been prepared for binding with lectins. The interaction events are marked by attachment of fluorescent dyes and gold
nanoparticles. The attachment of the fluorescent dyes and gold nanoparticles is achieved by standard avidin–biotin chemistry.
The detection principle is fluorescence or resonance light scattering (RLS). The electroless deposition of silver onto the
gold particles has been employed for RLS signal enhancement. Well-defined recognition systems, three monosaccharides (Man-α,
Glc-β, and Gal-β) or three glycoproteins (Asf, RNase A, and RNase B) with two lectins (ConA and RCA120), are chosen here to
establish the microarray-based assay, respectively. Highly selective recognition of carbohydrate–protein down to 25.6pg/mL
for RCA120 in solution and 8μM for Gal-β and 32ng/mL for Asf on the microarray spots is demonstrated.
Key wordsGlycan microarray-fluorescence-resonance light scattering-gold nanoparticle
10/2009: pages 145-153;
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ABSTRACT: Water-soluble supramolecular inclusion complexes of alpha-, beta-, and gamma-cyclodextrin-bicapped C60 (CD/C60) have been investigated for their photoinduced DNA cleavage activities, with the aim to assess the potential health risks of this class of compounds and to understand the effect of host cyclodextrins having different cavity dimensions. Factors such as incubation temperature, irradiation time, and concentration of NADH or CDs/ C60 supramolecular inclusion complexes have been examined. The results show that alpha-, beta-, and gamma-CDs/C60 are all able to cleave double-stranded DNA under visible light irradiation in the presence of NADH. However, a difference in the photoinduced DNA cleavage efficiency is observed, where the cleavage efficiency increases in the order of alpha-, beta-, and gamma-CD/C60. The difference is attributed to the different aggregation behavior of the inclusion complexes in aqueous solution, which is correlated to the cavity dimension of the host cyclodextrin molecules.
Environmental Science and Technology 09/2009; 43(15):5825-9. · 5.23 Impact Factor
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ABSTRACT: In this paper, a microarray-based surface-enhanced Raman spectroscopic (SERS) assay for detection of kinase functionality and inhibition has been reported. Biotinylated anti-phosphoserinen antibodies mark the phosphorylation and inhibition events and gold nanoparticles are attached to the antibodies by standard avidin-biotin chemistry, followed by silver deposition for SERS signal enhancement. The avidin conjugated fluorescein is used as SERS probe. The alpha-catalytic subunit of cyclic adenosine 5'-monophosphate (cAMP) dependent protein kinase (PKA), its well known substrate, kemptide, and three inhibitors, H89, HA1077, and KN62 have been chosen here to establish the SERS assay. As expected, highly selective inhibition of PKA is demonstrated with the inhibitor H89 and the inhibition assay enable to detect kinase inhibition as well as derive IC(50) (half maximal inhibitory concentration) plots.
Biosensors & bioelectronics 08/2009; 24(11):3335-9. · 5.43 Impact Factor
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ABSTRACT: Water-soluble supramolecular inclusion complexes of α-, β-, and γ-cyclodextrin-bicapped C60 (CD/C60) have been investigated for their photoinduced DNA cleavage activities, with the aim to assess the potential health risks of this class of compounds and to understand the effect of host cyclodextrins having different cavity dimensions. Factors such as incubation temperature, irradiation time, and concentration of NADH or CDs/C60 supramolecular inclusion complexes have been examined. The results show that α-, β-, and γ-CDs/C60 are all able to cleave double-stranded DNA under visible light irradiation in the presence of NADH. However, a difference in the photoinduced DNA cleavage efficiency is observed, where the cleavage efficiency increases in the order of α-, β-, and γ-CD/C60. The difference is attributed to the different aggregation behavior of the inclusion complexes in aqueous solution, which is correlated to the cavity dimension of the host cyclodextrin molecules.
07/2009;
