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Zhenxin Wang - One of the best experts on this subject based on the ideXlab platform.
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profiling of multiple matrix metalloproteinases activities in the progression of osteosarcoma by peptide microarray based Fluorescence Assay on polymer brush coated zinc oxide nanorod substrate
Sensors and Actuators B-chemical, 2021Co-Authors: Minghong Jian, Zhenxin Wang, Hua ZhangAbstract:Abstract Osteosarcoma (OS) is a primary malignant bone tumor with high rate of recurrence and lung metastasis. Matrix metalloproteinases (MMPs) are demonstrated as important biomarkers of tumor invasion and metastasis. Herein, a peptide microarray-based Fluorescence Assay is proposed to profile multiple MMPs (MMP-1, -2, -3, -7, -9 and -13) activities in the progression of OS by using the mouse-bearing xenograft U-2OS and Saos-2 human OSs. The peptide microarray containing different biotinylated peptide substrate spots is fabricated on the poly(glycidyl methacrylate-co-2-hydroxyethyl methacrylate) brush coated zinc oxide nanorod (ZnONR@P(GMA-HEMA) decorated glass slides, and labeled by cyanine 3 (Cy3) modified avidin (Avidin-Cy3) to produce Fluorescence signal. Special cleavage of peptide substrate by MMP resulted the decrease of Fluorescence signal. The MMPs activities are positively correlated with the change of Fluorescence intensity. This method has excellent selectivity and sensitivity, which enables to detect the activities of cellular secreted MMP-1, -2, -3, -7, -9, and -13 with limit of detection downs to 10 pmol L−1, 30 pmol L−1, 113 pmol L−1, 13 pmol L−1, 93 pmol L−1 and 12 pmol L−1, respectively. Furthermore, it is demonstrated that the activity pattern of MMPs in serum closely relevant to the disease progression and type of tumor.
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array based in situ Fluorescence Assay for profiling multiplex matrix metalloproteinases activities in tissue section
Analytica Chimica Acta, 2019Co-Authors: Zhen Lei, Minghong Jian, Jia Wei, Yaoqi Wang, Xianying Meng, Zhenxin WangAbstract:Herein, an array-based in situ Fluorescence Assay is proposed for high-throughput analysis and localization of multiplex matrix metalloproteinases (MMPs) activities in cell monolayers and tissue sections. Five specific MMPs (MMP-2, -3, -7, -9, and -14) peptide substrates containing FAM/Dabcyl fluorescent resonance energy transfer (FRET) pair are directly spotted on the surface of cell monolayers or tissue sections, and hydrolyzed by localized MMPs, resulting in Fluorescence recovery of FAM. MMPs activities are determined by the Fluorescence intensity of stained cells/tissues due to the cellular internalization of peptide fragments with FAM moiety. We demonstrate that the array-based in situ Fluorescence Assay is suitable for identifying the MMPs expression patterns of cells, as well as determining the secreted MMPs activities in cell monolayer with high sensitivity (as low as hundreds of cells per square centimeter). The feasibility of the Assay is further confirmed by evaluating inhibition potencies of six compounds toward five MMPs. Profiling of five MMPs activities in the localized parts of 32 thyroid tissues is performed without separation or extraction procedures, demonstrating the good practicality of the method.
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evaluation of matrix metalloproteinase inhibition by peptide microarray based Fluorescence Assay on polymer brush substrate and in vivo assessment
ACS Applied Materials & Interfaces, 2017Co-Authors: Zhen Lei, Yaoqi Wang, Xianying Meng, Hua Zhang, Hongda Chen, Zhenxin WangAbstract:Matrix metalloproteinases (MMPs) are important biomarkers and potential therapeutic targets of tumor. In this report, a peptide microarray-based Fluorescence Assay is developed for MMPs inhibitors evaluation through immobilization of biotin-modified peptides on the poly(glycidyl methacrylate-co-2-hydroxyethyl methacrylate) (P(GMA-HEMA)) brush-modified glass slides. After biotin is recognized with cyanine 3 (Cy3)-modified avidin (Cy3-avidin), the microarrays can produce strong Fluorescence signal. The biotin moieties detach from microarray, when the biotin-modified peptide substrates are specially cleaved by a MMP, resulting in decreased Fluorescence intensity of the microarray. The decreasing level of Fluorescence intensity is correlated with the MMP inhibition. Nine known MMP inhibitors against MMP-2 and MMP-9 are evaluated by the Assay, and the quantitative determination of inhibitory potencies (half maximal inhibitory concentration) are obtained, which are comparable with the literatures. Two biocompatible fluorogenic peptides containing MMP-specific recognition sequences and FAM/Dabcyl fluorophore-quencher pair are designed as activatable reporter probes for sensing MMP-2 and MMP-9 activities in cell and in vivo. The peptide microarray-based results are well verified by the cell inhibition Assay and in vitro Fluorescence imaging, and further confirmed by the in vivo imaging of HT-1080 tumor-bearing mice.
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peptide microarray based metal enhanced Fluorescence Assay for multiple profiling of matrix metalloproteinases activities
Analytical Chemistry, 2017Co-Authors: Zhen Lei, Yaoqi Wang, Xianying Meng, Hua Zhang, Zhenxin WangAbstract:Matrix metalloproteinases (MMPs) are closely associated with cancer cell invasion and metastasis. Herein, a Fluorescence resonance energy transfer (FRET)-peptide microarray-based metal enhanced Fluorescence (MEF) Assay is proposed for multiple and sensitive profiling of MMPs activities on a novel Au/Ag@SiO2 substrate. The Au/Ag@SiO2 substrate is prepared by electroless deposition of silver on gold nanoparticle (GNP) seeds, followed by SiO2 shell coating and surface functionalization. The specific FRET peptides are spotted on the Au/Ag@SiO2 substrate to sensitively detect MMPs (MMP-2, -3, -7, -9, -14) via Fluorescence recovery by the MMP cleavage of quenched peptide motifs and further enhanced by MEF. Under the optimal conditions, the limits of detection are 12.2 fg mL–1 for MMP-2, 60 pg mL–1 for MMP-3, 0.22 pg mL–1 for MMP-7, 102 fg mL–1 for MMP-9, and 0.68 ng mL–1 for MMP-14, respectively. The practicability of the FRET-peptide microarray-based MEF Assay is demonstrated by profiling of multiplexed MMPs a...
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sensitive detection of polynucleotide kinase activity by paper based Fluorescence Assay with λ exonuclease assistance
Analytical Chemistry, 2016Co-Authors: Hua Zhang, Zhen Lei, Zhen Zhao, Zhenxin WangAbstract:The phosphorylation of nucleic acid with 5′-OH termini catalyzed by polynucleotide kinase (PNK) involves several significant cellular events. Here a paper-based Fluorescence Assay with λ exonuclease assistance was reported for facile detection of PNK activity through monitoring the change of Fluorescence intensity on paper surface. Cy5-labeled ssDNA was first immobilized on the surface of aldehyde group modified paper, and BHQ-labeled ssDNA was then employed to quench the Fluorescence of the immobilized Cy5-labeled ssDNA with the help of an adaptor ssDNA. When PNK and λ exonuclease cleavage reaction were introduced, the Fluorescence quenching effect on the paper surface was blocked because of the digestion of phosphorylated dsDNA by the coupled enzymes. By using this paper-based Assay, PNK activity both in pure reaction buffer and in practical biosample have been successfully measured. Highly sensitive detection of PNK activity down to 0.0001 U mL–1 and lysate of about 50 cells is achieved. The inhibition...
Jianhui Jiang - One of the best experts on this subject based on the ideXlab platform.
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a novel Fluorescence Assay for inorganic pyrophosphatase based on modulated aggregation of graphene quantum dots
Analyst, 2016Co-Authors: Xueli Zhu, Jinwen Liu, Haiyang Peng, Jianhui JiangAbstract:A simple and highly sensitive fluorometric method has been developed for inorganic pyrophosphatase (PPase) activity detection based on the disaggregation and aggregation of graphene quantum dots (GQDs). Copper ions can trigger the severe aggregation of GQDs with rich carboxyl groups, which results in effective Fluorescence quenching. While, with the addition of pyrophosphate (PPi), the quenched Fluorescence is effectively recovered owing to the strong interaction between PPi and Cu2+. Furthermore, under the catalytic hydrolysis of PPase, the complex of PPi–Cu2+–PPi is rapidly disassembled, and the Fluorescence is re-quenched. This method is highly sensitive and selective for PPase detection, with a linear correlation between the Fluorescence intensity and the PPase concentration in the range from 1 to 200 mU mL−1 with a detection limit down to 1 mU mL−1 (S/N = 3). Additionally, the inhibition effect of NaF on the PPase activity is also studied. Thus, the proposed method may hold a potential application in the diagnosis of PPase-related diseases and screening of PPase inhibitors, to evaluate the function and inhibition of PPase in biological systems.
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a sensitive homogeneous Fluorescence Assay for detection of thymine dna glycosylase activity based on exonuclease mediated amplification
Chemical Communications, 2013Co-Authors: Cuihua Chen, Manfen Liang, Dianming Zhou, Hao Tang, Jianhui JiangAbstract:A novel homogeneous Fluorescence Assay strategy for highly sensitive detection of thymine DNA glycosylase (TDG) enzyme activity based on the exonuclease-mediated signal amplification reaction was reported.
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a label free exonuclease iii aided Fluorescence Assay for adenosine triphosphate based on graphene oxide and ligation reaction
New Journal of Chemistry, 2013Co-Authors: Wenping Zhu, Jianhui Jiang, Ziwei Zhao, Guoli ShenAbstract:A label free exonuclease III (Exo III)-aided Fluorescence Assay for adenosine triphosphate (ATP) was developed based on the ATP-dependent enzymatic reaction and graphene oxide (GO). This strategy relies on the principle that Exo III shows different cleavage capacity for a DNA substrate in the absence and presence of ATP and the preferential binding of GO to single-stranded DNA over double-stranded one. By combining the unique properties of SYBR Green I and GO adsorption, this sensor displays an improved sensitivity and a wide linear range within the ATP concentration from 1 nM to 200 nM with a low detection limit of 0.2 nM. The proposed method is simple, cost-effective and convenient, which might create a new methodology for developing a sensitive ATP biosensor.
Zhen Lei - One of the best experts on this subject based on the ideXlab platform.
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array based in situ Fluorescence Assay for profiling multiplex matrix metalloproteinases activities in tissue section
Analytica Chimica Acta, 2019Co-Authors: Zhen Lei, Minghong Jian, Jia Wei, Yaoqi Wang, Xianying Meng, Zhenxin WangAbstract:Herein, an array-based in situ Fluorescence Assay is proposed for high-throughput analysis and localization of multiplex matrix metalloproteinases (MMPs) activities in cell monolayers and tissue sections. Five specific MMPs (MMP-2, -3, -7, -9, and -14) peptide substrates containing FAM/Dabcyl fluorescent resonance energy transfer (FRET) pair are directly spotted on the surface of cell monolayers or tissue sections, and hydrolyzed by localized MMPs, resulting in Fluorescence recovery of FAM. MMPs activities are determined by the Fluorescence intensity of stained cells/tissues due to the cellular internalization of peptide fragments with FAM moiety. We demonstrate that the array-based in situ Fluorescence Assay is suitable for identifying the MMPs expression patterns of cells, as well as determining the secreted MMPs activities in cell monolayer with high sensitivity (as low as hundreds of cells per square centimeter). The feasibility of the Assay is further confirmed by evaluating inhibition potencies of six compounds toward five MMPs. Profiling of five MMPs activities in the localized parts of 32 thyroid tissues is performed without separation or extraction procedures, demonstrating the good practicality of the method.
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evaluation of matrix metalloproteinase inhibition by peptide microarray based Fluorescence Assay on polymer brush substrate and in vivo assessment
ACS Applied Materials & Interfaces, 2017Co-Authors: Zhen Lei, Yaoqi Wang, Xianying Meng, Hua Zhang, Hongda Chen, Zhenxin WangAbstract:Matrix metalloproteinases (MMPs) are important biomarkers and potential therapeutic targets of tumor. In this report, a peptide microarray-based Fluorescence Assay is developed for MMPs inhibitors evaluation through immobilization of biotin-modified peptides on the poly(glycidyl methacrylate-co-2-hydroxyethyl methacrylate) (P(GMA-HEMA)) brush-modified glass slides. After biotin is recognized with cyanine 3 (Cy3)-modified avidin (Cy3-avidin), the microarrays can produce strong Fluorescence signal. The biotin moieties detach from microarray, when the biotin-modified peptide substrates are specially cleaved by a MMP, resulting in decreased Fluorescence intensity of the microarray. The decreasing level of Fluorescence intensity is correlated with the MMP inhibition. Nine known MMP inhibitors against MMP-2 and MMP-9 are evaluated by the Assay, and the quantitative determination of inhibitory potencies (half maximal inhibitory concentration) are obtained, which are comparable with the literatures. Two biocompatible fluorogenic peptides containing MMP-specific recognition sequences and FAM/Dabcyl fluorophore-quencher pair are designed as activatable reporter probes for sensing MMP-2 and MMP-9 activities in cell and in vivo. The peptide microarray-based results are well verified by the cell inhibition Assay and in vitro Fluorescence imaging, and further confirmed by the in vivo imaging of HT-1080 tumor-bearing mice.
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peptide microarray based metal enhanced Fluorescence Assay for multiple profiling of matrix metalloproteinases activities
Analytical Chemistry, 2017Co-Authors: Zhen Lei, Yaoqi Wang, Xianying Meng, Hua Zhang, Zhenxin WangAbstract:Matrix metalloproteinases (MMPs) are closely associated with cancer cell invasion and metastasis. Herein, a Fluorescence resonance energy transfer (FRET)-peptide microarray-based metal enhanced Fluorescence (MEF) Assay is proposed for multiple and sensitive profiling of MMPs activities on a novel Au/Ag@SiO2 substrate. The Au/Ag@SiO2 substrate is prepared by electroless deposition of silver on gold nanoparticle (GNP) seeds, followed by SiO2 shell coating and surface functionalization. The specific FRET peptides are spotted on the Au/Ag@SiO2 substrate to sensitively detect MMPs (MMP-2, -3, -7, -9, -14) via Fluorescence recovery by the MMP cleavage of quenched peptide motifs and further enhanced by MEF. Under the optimal conditions, the limits of detection are 12.2 fg mL–1 for MMP-2, 60 pg mL–1 for MMP-3, 0.22 pg mL–1 for MMP-7, 102 fg mL–1 for MMP-9, and 0.68 ng mL–1 for MMP-14, respectively. The practicability of the FRET-peptide microarray-based MEF Assay is demonstrated by profiling of multiplexed MMPs a...
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sensitive detection of polynucleotide kinase activity by paper based Fluorescence Assay with λ exonuclease assistance
Analytical Chemistry, 2016Co-Authors: Hua Zhang, Zhen Lei, Zhen Zhao, Zhenxin WangAbstract:The phosphorylation of nucleic acid with 5′-OH termini catalyzed by polynucleotide kinase (PNK) involves several significant cellular events. Here a paper-based Fluorescence Assay with λ exonuclease assistance was reported for facile detection of PNK activity through monitoring the change of Fluorescence intensity on paper surface. Cy5-labeled ssDNA was first immobilized on the surface of aldehyde group modified paper, and BHQ-labeled ssDNA was then employed to quench the Fluorescence of the immobilized Cy5-labeled ssDNA with the help of an adaptor ssDNA. When PNK and λ exonuclease cleavage reaction were introduced, the Fluorescence quenching effect on the paper surface was blocked because of the digestion of phosphorylated dsDNA by the coupled enzymes. By using this paper-based Assay, PNK activity both in pure reaction buffer and in practical biosample have been successfully measured. Highly sensitive detection of PNK activity down to 0.0001 U mL–1 and lysate of about 50 cells is achieved. The inhibition...
Kirk S Schanze - One of the best experts on this subject based on the ideXlab platform.
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conjugated polyelectrolyte based real time Fluorescence Assay for adenylate kinase
Analytical Chemistry, 2009Co-Authors: Yan Liu, Kirk S SchanzeAbstract:Addition of adenosine 5'-triphosphate (ATP) to a solution of the anionic conjugated polyelectrolyte PPECO2 and copper(II) ion (Cu2+) recovers the Cu2+-quenched Fluorescence of PPECO(2) to a significantly greater extent compared with the addition of adenosine 5'-diphosphate (ADP) or adenosine 5'-monophosphate (AMP) at the same concentration levels. Taking advantage of the differential response of the PPECO2-Cu2+ system to ATP, ADP and AMP, we have developed Fluorescence turn-off and turn-on Assays that monitor the catalytic activity of adenylate kinase (ADK) in the equilibrium transphosphorylation reaction (ATP + AMP 2ADP). The Fluorescence turn-on and turn-off Assays monitor the forward and reverse transphosphorylation reactions, respectively. The forward Assay operates with ATP substrate present at the submillimolar concentration range and offers a straightforward and rapid detection of ADK catalytic activity with the enzyme present in the nanomolar range, in either end-point or real-time formats. The real-time Fluorescence intensity from PPECO2 can be converted to substrate (ATP) concentration in the forward reaction Assay by using an ex-situ calibration curve, allowing ADK catalyzed reaction rates and kinetic parameters to be determined. ADK activation by Mg2+ and inhibition by Ag+ and product are analyzed using the optimized Assay system. Non-specific interactions are observed between the Assay complex and other proteins, but the signal response to the ADK Assay is demonstrated to mainly arise from the specific enzyme catalyzed transphosphorylation reaction.
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conjugated polyelectrolyte based real time Fluorescence Assay for alkaline phosphatase with pyrophosphate as substrate
Analytical Chemistry, 2008Co-Authors: Yan Liu, Kirk S SchanzeAbstract:The Fluorescence of the anionic, carboxylate-substituted poly(phenylene ethynylene) polymer PPECO2 is quenched very efficiently via the addition of 1 equiv of Cu2+. Addition of pyrophosphate (PPi) into the weakly fluorescent solution of PPECO2 and Cu2+ induces recovery of the polymer’s Fluorescence; the recovery occurs because PPi complexes with Cu2+, effectively sequestering the ion so it cannot bind to the carboxylate groups of the polymer. A calibration curve was developed that relates the extent of Fluorescence recovery to [PPi], making the PPECO2−Cu2+ system a sensitive and selective turn-on sensor for PPi. Using the PPECO2−Cu2+ system as the signal transducer, a real-time Fluorescence turn-off Assay for the enzyme alkaline phosphatase (ALP) using PPi as the substrate is developed. The Assay operates with [PPi] in the micromolar range, and it offers a straightforward and rapid detection of ALP activity with the enzyme present in the nanomolar concentration range, operating either in an end point or r...
Xiaojun Zhang - One of the best experts on this subject based on the ideXlab platform.
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one step ultrasonic synthesis of graphene quantum dots with high quantum yield and their application in sensing alkaline phosphatase
Chemical Communications, 2015Co-Authors: Guangfeng Wang, Hong Jiang, Ling Chen, Xiaojun ZhangAbstract:With only graphene oxide and KMnO4, the luminescent graphene quantum dots (GQDs) in high quantum yield were prepared by one-step synthesis using ultrasonication, and applied in the label-free, simple and fast Fluorescence Assay of alkaline phosphatase (ALP).