He Zhang

Hunan Institute of Engineering, Siangtan, Hunan, China

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Publications (8)34.32 Total impact

  • He Zhang, Xinjiang Hu, Xin Fu
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    ABSTRACT: This study reports the development of an aptamer-mediated microfluidic beads-based sensor for multiple analytes detection and quantification using multienzyme-linked nanoparticle amplification and quantum dots labels. Adenosine and cocaine were selected as the model analytes to validate the assay design based on strand displacement induced by target–aptamer complex. Microbeads functionalized with the aptamers and modified electron rich proteins were arrayed within a microfluidic channel and were connected with the horseradish peroxidases (HRP) and capture DNA probe derivative gold nanoparticles (AuNPs) via hybridization. The conformational transition of aptamer induced by target–aptamer complex contributes to the displacement of functionalized AuNPs and decreases the fluorescence signal of microbeads. In this approach, increased binding events of HRP on each nanosphere and enhanced mass transport capability inherent from microfluidics are integrated for enhancing the detection sensitivity of analytes. Based on the dual signal amplification strategy, the developed aptamer-based microfluidic bead array sensor could discriminate as low as 0.1 pM of adenosine and 0.5 pM cocaine, and showed a 500-fold increase in detection limit of adenosine compared to the off-chip test. The results proved the microfluidic-based method was a rapid and efficient system for aptamer-based targets assays (adenosine (0.1 pM) and cocaine (0.5 pM)), requiring only minimal (microliter) reagent use. This work demonstrated the successful application of aptamer-based microfluidic beads array sensor for detection of important molecules in biomedical fields.
    Biosensors & Bioelectronics 01/2014; 57:22–29. · 6.45 Impact Factor
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    Xin Fu, Yuan Liu, Zhitao Wu, He Zhang
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    ABSTRACT: A simple, highly sensitive and selective method based on the rhodamine B-covered gold nanoparticle with dual-readout (colorimetric and fluorometric) detection for -cysteine is proposed. A mechanism is that citrate-stabilized AuNPs were modified with RB by electrostatic interaction, which enables the nanometal surface energy transfer (NSET) from the RB to the AuNPs, quenching the fluorescence. In the presence of -cysteine, it was used as a competitor in the NSET by the strongly Au-S bonding to release RB from the Au surface and recover the fluorescence, and the red-to-purple color change quickly, which was monitored simply by the naked eye. Under the optimum conditions, the detection limit is as low as 10 nM. The method possessed the advantages of simplicity, rapidity and sensitivity at the same time. The method was also successfully applied to the determination of -cysteine in human urine samples, and the results were satisfying.
    Bulletin- Korean Chemical Society 01/2014; 35(4). · 0.84 Impact Factor
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    ABSTRACT: This study reports the development of a microfluidic bead-based nucleic acid sensor for sensitive detection of circulating tumor cells in blood samples using multienzyme-nanoparticle amplification and quantum dot labels. In this method, the microbeads functionalized with the capture probes and modified electron rich proteins were arrayed within a microfluidic channel as sensing elements, and the gold nanoparticles (AuNPs) functionalized with the horseradish peroxidases (HRP) and DNA probes were used as labels. Hence, two signal amplification approaches are integrated for enhancing the detection sensitivity of circulating tumor cells. First, the large surface area of Au nanoparticle carrier allows several binding events of HRP on each nanosphere. Second, enhanced mass transport capability inherent from microfluidics leads to higher capture efficiency of targets because continuous flow within micro-channel delivers fresh analyte solution to the reaction site which maintains a high concentration gradient differential to enhance mass transport. Based on the dual signal amplification strategy, the developed microfluidic bead-based nucleic acid sensor could discriminate as low as 5fM (signal-to-noise (S/N)3) of synthesized carcinoembryonic antigen (CEA) gene fragments and showed a 1000-fold increase in detection limit compared to the off-chip test. In addition, using spiked colorectal cancer cell lines (HT29) in the blood as a model system, the detection limit of this chip-based approach was found to be as low as 1 HT29 in 1mL blood sample. This microfluidic bead-based nucleic acid sensor is a promising platform for disease-related nucleic acid molecules at the lowest level at their earliest incidence.
    Analytica chimica acta 05/2013; 779:64-71. · 4.31 Impact Factor
  • He ZHANG, Xin FU, Zhen-Jun ZHU
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    ABSTRACT: A novel approach for single-nucleotide detection based on the micropump-integrated microfluidic microbeads array chip and an apyrase-mediated primer extension process was developed. In this method, an integrated chip was constructed by a microfluidic chip, a primers-modified microbeads array and a micro-pump driven by evaporation and capillary effect. The target DNA flowed across the fabricated microfluidic beads array and hybridized with immobilized primer sequences. When the 3′ terminus of primer matched with the target DNA in the single-base mutation site of interest, under synergistic effect of apyrase and exonuclease-deficient Klenow DNA polymerase, the matched primer extended along the template DNA sequence and incorporated the biotin-dCTP into the extended primers and immobilized it onto the surface of microbeads. Then, the streptavidin-labeled quantum dots bond with deposited biotin moities of biotin-dCTP and generated a fluorescence signal. On the contrary, there is no signal when signal-base mismatched duplexes were present in the 3′ terminus of the primer. The limit of detection is 0.2 pM target DNA (S/N > 3) for micro-pump driven chip and 0.5 pM target DNA for liquid pressure driven chip respectively. The chip-based signal enhancement for single-nucleotide discrimination using micro-pump integrated microfluidic chip resulted in 500 times higher sensitivity than that of an off-chip test. Since the off-chip assay only detected 0.1 nM target DNA. The fluorescence signals are linear in the target DNA concentration ranging 0.5 pM to 30 pM. This method was also used to detect two multi-drug resistance gene 1 (MDR1)-associated SNP sites (C3435T and G2677T) from a human genomic sample. The fluorescence signals indicated the subject used here possessed both MDR1 3435CT and MDR1 2677TT genotypes, which were consistent with the results by DNA sequencing. This approach displays good specificity, sensitivity and stability for discrimination of gene mutation.
    Chinese Journal of Analytical Chemistry 04/2013; 41(4):473–480. · 0.79 Impact Factor
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    ABSTRACT: This study reports the development of a microfluidic beads-based immunosensor for sensitive detection of cancer biomarker α-fetoprotein (AFP) that uses multienzyme-nanoparticle amplification and quantum dots labels. This method utilizes microbeads functionalized with the capture antibodies (Ab(1)) and modified electron rich proteins as sensing platform that was fabricated within a microfluidic channel, and uses gold nanoparticles (AuNPs) functionalized with the horseradish peroxidase (HRP) and the detection antibodies (Ab(2)) as label. Greatly enhanced sensitivity for the cancer biomarker is based on a dual signal amplification strategy: first, the large surface area of Au nanoparticle carrier allows several binding events of HRP on each nanosphere. Enhanced sensitivity was achieved by introducing the multi-HRP-antibody functionalized AuNPs onto the surface of microbeads through "sandwich" immunoreactions and subsequently multiple biotin moieties could be deposited onto the surface of beads resulted from the oxidation of biotin-tyramine by hydrogen peroxide. Streptavidin-labeled quantum dots were then allowed to bind to the deposited biotin moieties and displayed the signal. Secondly, enhanced mass transport capability inherent from microfluidics leads to higher capture efficiency of targets because continuous flow within micro-channel delivers fresh analyte solution to the reaction site which maintains a high concentration gradient differential to enhance mass transport. Based on the dual signal amplification strategy, the developed microfluidic bead-based immunosensor could discriminate as low as 0.2pgmL(-1) AFP in 10μL of undiluted calf serum (0.2fg/chip), and showed a 500-fold increase in detection limit compared to the off-chip test and 50-fold increase in detection limit compared to microfluidic beads-based immunoassay using single label HRP-Ab(2). The immunosensor showed acceptable repeatability and reproducibility. This microfluidic beads-based immunosensor is a promising platform for disease-related biomolecules at the lowest level at their earliest incidence.
    Biosensors & Bioelectronics 11/2012; 42C:23-30. · 6.45 Impact Factor
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    ABSTRACT: This study reports the development of an on-chip enzyme-mediated primer extension process based on a microfluidic device with microbeads array for single-nucleotide discrimination using quantum dots as labels. The functionalized microbeads were independently introduced into the arrayed chambers using the loading chip slab. A single channel was used to generate weir structures to confine the microbeads and make the beads array accessible by microfluidics. The applied allele-specific primer extension method employed a nucleotide-degrading enzyme (apyrase) to achieve specific single-nucleotide detection. Based on the apyrase-mediated allele-specific primer extension with quantum dots as labels, on-chip single-nucleotide discrimination was demonstrated with high discrimination specificity and sensitivity (0.5 pM, signal/noise > 3) using synthesized target DNA. The chip-based signal enhancement for single-nucleotide discrimination resulted in 200 times higher sensitivity than that of an off-chip test. This microfluidic device successfully achieved simultaneous detection of two disease-associated single-nucleotide polymorphism sites using polymerase chain reaction products as target. This apyrase-mediated microfluidic primer extension approach combines the rapid binding kinetics of homogeneous assays of suspended microbeads array, the liquid handling capability of microfluidics, and the fluorescence detection sensitivity of quantum dots to provide a platform for single-base analysis with small reagent consumption, short assay time, and parallel detection.
    Analytical Biochemistry 04/2012; 426(1):30-9. · 2.58 Impact Factor
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    ABSTRACT: A novel microfluidic device with microbeads array was developed and sensitive genotyping of human papillomavirus was demonstrated using a multiple-enzyme labeled oligonucleotide-Au nanoparticle bioconjugate as the detection tool. This method utilizes microbeads as sensing platform that was functionalized with the capture probes and modified electron rich proteins, and uses the horseradish peroxidase (HRP)-functionalized gold nanoparticles as label with a secondary DNA probe. The functionalized microbeads were independently introduced into the arrayed chambers using the loading chip slab. A single channel was used to generate weir structures to confine the microbeads and make the beads array accessible by microfluidics. Through "sandwich" hybridization, the enzyme-functionalized Au nanoparticles labels were brought close to the surface of microbeads. The oxidation of biotin-tyramine by hydrogen peroxide resulted in the deposition of multiple biotin moieties onto the surface of beads. This deposition is markedly increased in the presence of immobilized electron rich proteins. Streptavidin-labeled quantum dots were then allowed to bind to the deposited biotin moieties and displayed the signal. Enhanced detection sensitivity was achieved where the large surface area of Au nanoparticle carriers increased the amount HRP bound per sandwiched hybridization. The on-chip genotyping method could discriminate as low as 1fmol/L (10zmol/chip, SNR>3) synthesized HPV oligonucleotides DNA. The chip-based signal enhancement of the amplified assay resulted in 1000 times higher sensitivity than that of off-chip test. In addition, this on-chip format could discriminate and genotype 10copies/μL HPV genomic DNA using the PCR products. These results demonstrated that this on-chip approach can achieve highly sensitive detection and genotyping of target DNA and can be further developed for detection of disease-related biomolecules at the lowest level at their earliest incidence.
    Biosensors & Bioelectronics 08/2011; 29(1):89-96. · 6.45 Impact Factor
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    ABSTRACT: In this study, a novel microfluidic device with microbead array was developed and sensitive genotyping of HBV was demonstrated using quantum dot as labels. This device was assembled by using two PDMS slabs featured with different microstructures and channel depths for the construction of a functional region comprising a chamber array and a single sampling microchannel. Since the chamber array and its sampling channel are of different channel depths and are bonded face-to-face, weir structures are generated to confine the microbeads which could be addressed using the microfluidic channel. Highly sensitive virus DNA detection was achieved by the enhanced mass transport in the microfluidics and the rapid reaction dynamics of suspension microbead array. The device could detect 1000 copies/mL of HBV virus in clinical serum samples using in vitro transcribed RNA as the target molecules. Based on DNA hybridization with quantum dots labels, on-chip virus genotyping was also demonstrated with high discrimination specificity and sensitivity (4 pM, S/N >3) using synthesized HBV DNA probes. This microfluidic device combines the rapid binding kinetics of homogeneous assays of microbead array, the liquid handling capability of microfluidics, and the fluorescence detection sensitivity of quantum dots to provide a platform for high sensitivity virus DNA analysis with small reagent consumption, short assay time and parallel detection.
    Biosensors & Bioelectronics 07/2010; 25(11):2402-7. · 6.45 Impact Factor