Browsing by Author "Chuang, Yao-Chen"

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A method of layer-by-layer gold nanoparticle hybridization in a quartz crystal microbalance DNA sensing system used to detect dengue virus
(2009-05-27)
Chen, Sz-Hau
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Chuang, Yao-Chen
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Lu, Yi-Chen
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Lin, Hsiu-Chao
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Yang, Yun-Liang
Dengue virus (DENV) is nowadays the most important arthropod-spread virus affecting humans existing in more than 100 countries worldwide. A rapid and sensitive detection method for the early diagnosis of infectious dengue virus urgently needs to be developed. In the present study, a circulating-flow quartz crystal microbalance (QCM) biosensing method combining oligonucleotide-functionalized gold nanoparticles (i.e. AuNP probes) used to detect DENV has been established. In the DNA-QCM method, two kinds of specific AuNP probes were linked by the target sequences onto the QCM chip to amplify the detection signal, i.e. oscillatory frequency change (Delta F) of the QCM sensor. The target sequences amplified from the DENV genome act as a bridge for the layer-by-layer AuNP probes' hybridization in the method. Besides being amplifiers of the detection signal, the specific AuNP probes used in the DNA-QCM method also play the role of verifiers to specifically recognize their target sequences in the detection. The effect of four AuNP sizes on the layer-by-layer hybridization has been evaluated and it is found that 13 nm AuNPs collocated with 13 nm AuNPs showed the best hybridization efficiency. According to the nanoparticle application, the DNA-QCM biosensing method was able to detect dengue viral RNA in virus-contaminated serum as plaque titers being 2 PFU ml(-1) and a linear correlation (R(2) = 0.987) of Delta F versus virus titration from 2 x 10(0) to 2 x 10(6) PFU ml(-1) was found. The sensitivity and specificity of the present DNA-QCM method with nanoparticle technology showed it to be comparable to the fluorescent real-time PCR methods. Moreover, the method described herein was shown to not require expensive equipment, was label-free and highly sensitive.
An optical biosensing platform for proteinase activity using gold nanoparticles
(2010-08-01)
Chuang, Yao-Chen
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Li, Jung-Chun
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Chen, Sz-Hau
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Liu, Ting-Yu
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Kuo, Ching-Han
The surface plasmon resonance (SPR) wavelength of colloidal gold nanoparticles (AuNPs) can vary when the AuNPs aggregate, have different sizes or shapes, or are modified with chemical molecules. In this study, an optical biosensing platform for a proteinase activity assay was established based on the SPR property of AuNPs. The 13-nm AuNPs were modified with gelatin (AuNPs-gelatin) as a proteinase substrate and subsequently modified with 6-mercaptohexan-1-ol (MCH) (AuNPs/MCH-gelatin). After proteinase (trypsin or gelatinase) digestion, the AuNPs lose shelter, and MCH increases the attractive force between the modified AuNPs. Therefore, the AuNPs gradually move closer to each other, resulting in AuNPs aggregation. The AuNPs aggregation can be monitored by the red shift of surface plasmon absorption and a visible color change of the AuNPs is from red to blue. Such a color change can be observed with the naked eye. For detection, the absorption ratio, A(625)/A(525), of the reacted AuNPs solution can be used to estimate quantitatively the proteinase activity. A linear correlation has been established with trypsin activity at concentrations from 1.25 x 10(-1) to 1.25 x 10(2) U and matrix metalloproteinase-2 activity at concentrations from 50 ng/mL to 600 ng/mL. (C) 2010 Elsevier Ltd. All rights reserved.
Aqueous zymography screening of matrix metalloproteinase activity and inhibition based on colorimetric gold nanoparticles
(2012-02-15)
Chuang, Yao-Chen
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Huang, Wei-Ting
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Chiang, Pin-Hsuan
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Tang, Meng-Che
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Lin, Chih-Sheng
An optical gold nanoparticles (AuNPs)-based method was fabricated for the rapid detection of matrix metalloproteinase (MMP) activity and screening potential MMP inhibitors without sophisticated instruments. The diagnosis platform was composed of AuNPs, particular MMP substrates and 6-mercapto-1-hexanol (MCH). The functionalized AuNPs were subjected to specific MMP digestion, and the MMP found the substrate on AuNPs, such that the AuNPs lost shelter and MCH increased the attraction force between AuNPs. Consequently, AuNPs aggregation and a color change from red to purple with increasing MMP concentration were observed. The surface plasmon resonance (SPR) of the formed AuNPs allowed for the quantitative detection of MMP activity. A sensitive linear correlation existed between the absorbance and the activity of the MMPs, which ranged from 10 ng/mL to 700 ng/mL in NTTC buffer and plasma samples. The proposed colorimetric method could be accomplished in a homogeneous solution with one-step operation in 30 min and has been successfully applied to the determination of particular MMP activity in plasma samples, in which the results are consistent with substrate zymography. This technology may become a simple platform for parallel screening a number of inhibitors and offer an alternative method to studying the efficiency of inhibitors for suppressing MMP activity. The absorbance ratio at 625 nm and 525 nm (A(625)/A(525)) confirmed the efficiency of the inhibitors as observed in substrate zymography. The IC50 of ONO-4817 and galardin for MMP-1, MMP-2 and MMP-7 determined by the proposed colorimetric method was similar to the results of substrate zymography. Crown Copyright (C) 2011 Published by Elsevier B.V. All rights reserved.
Disposable amperometric immunosensing strips fabricated by Au nanoparticles-modified screen-printed carbon electrodes for the detection of foodborne pathogen Escherichia coli O157 : H7
(2008-07-15)
Lin, Yueh-Hui
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Chen, Sz-Hau
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Chuang, Yao-Chen
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Lu, Yi-Chen
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Shen, Thomas Y.
A disposable amperometric immunosensing strip was fabricated for rapid detection of Escherichia coli O157:H7. The method uses an indirect sandwich enzyme-linked immunoassay with double antibodies. Screen-printed carbon electrodes (SPCEs) were framed by commercial silver and carbon inks. For electrochemical characterization the carbon electrodes were coupled with the first E. coli O157:H7-specific antibody, E. coli O157:H7 intact cells and the second E. coli O157:H7-specific antibody conjugated with horseradish peroxidase (HRP). Hydrogen peroxide and ferrocenedicarboxylic acid (FeDC) were used as the substrate for HRP and mediator, respectively, at a potential +300 mV vs. counter/reference electrode. The response current (RC) of the immunosensing strips could be amplified significantly by 13-nm diameter Au nanoparticles (AuNPs) attached to the working electrode. The results show that the combined effects of AuNPs and FeDC enhanced RC by 13.1-fold. The SPCE immunosensing strips were used to detect E. coli O157:H7 specifically. Concentrations of E. coli O157:H7 from 10(2) to 10(7) CFU/ml could be detected. The detection limit was approximately 6 CFU/strip in PBS buffer and 50 CFU/strip in milk. The SPCE modified with AuNPs and FeDC has the potential for further applications and provides the basis for incorporating the method into an integrated system for rapid pathogen detection. (c) 2008 Elsevier B.V. All rights reserved.
Dual functional AuNRs@MnMEIOs nanoclusters for magnetic resonance imaging and photothermal therapy
(2014-05-01)
Chuang, Yao-Chen
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Lin, Chia-Jung
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Lo, Shih-Feng
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Wang, Jei-Lin
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Tzou, Shey-Cherng
A novel dual functional theranosis platform is developed based on manganese magnetism-engineered iron oxide (MnMEIO) and gold nanorods (AuNRs) to combine magnetic resonance (MR) imaging and photothermal therapy in one nanocluster. The platform showed improved T-2-weighted MR imaging and exhibited a near-infrared (NIR) induced temperature elevation due to the unique characteristics of AuNRs@MnMEIOs nanoclusters. The obtained dual functional spherical-shaped nanoclusters showed low cytotoxicity, and high cellular uptake efficiency. The AuNRs@MnMEIOs nanoclusters also demonstrated a 1.9 and 2.2 folds r(2) relaxivity value higher than those of monodispersed MnMEIO and Resovist. In addition, in vivo MR imaging study found that the contrast enhancements were -70.4 +/- 4.3% versus 7.5 +/- 3.0% in Her-2/neu overexpression tumors as compared to the control tumors. More importantly, NIR laser irradiation to the tumor site resulted in outstanding photothermal therapeutic efficacy and without damage to the surrounding tissue. In additional, the prepared dual functional AuNRs@MnMEIOs display high stability and furthermore disperse even in the presence of external magnet, showing that AuNRs@MnMEIOs nanoclusters can be manipulated by an external magnetic field. Therefore, such nanoclusters combined MR imaging and photothermal therapeutic functionality can be developed as a promising nanosystem for effective cancer diagnosis and therapy. (C) 2014 Elsevier Ltd. All rights reserved.
Optical Detection of Human Papillomavirus Type 16 and Type 18 by Sequence Sandwich Hybridization With Oligonucleotide-Functionalized Au Nanoparticles
(2009-06-01)
Chen, Sz-Hau
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Lin, Kun-I
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Tang, Chuan-Yi
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Peng, Sheng-Lung
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Chuang, Yao-Chen
The importance of detecting and subtyping human papillomaviruses (HPVs) in clinical and epidemiological studies has been well addressed. In detecting the most common types of HPV, type 16 (HPV-16) and type 18 (HPV-18), in the cervical mucous of patients in a simple and rapid manner, the assay of a label-free colorimetric DNA sensing method based on sequence sandwich hybridization with oligonucleotide-functionalized Au nanoparticles (AuNPs) was fabricated in this study. Specific oligonucleotide probes were designed for the sequence detection within the L1 gene of HPV-16 and HPV-18, and the probes were capped onto AuNPs, as AuNP probes. The target HPV sequences in clinical specimens were obtained by an asymmetric polymerase chain reaction (PCR) with universal primers, which can amplify the target sequences from several HPV serotypes, including HPV-16 and HPV-18. The DNA sandwich hybridization between the target sequences and the specific AuNP probes was performed at a temperature closer to the theoretical melting temperature of the DNA hybridization. Next, the procedure of increasing salt concentration and cooling the hybridizing solution was immediately utilized to discriminate the target sequences of HPV-16 or HPV-18. If the target sequences were not complementary to sequences of AuNP probes, the AuNPs would aggregate because no duplex DNA formation occurred such that the color of the reaction solution changed from red to purple. If the AuNP probes were a perfect match to the target sequences and a full DNA sandwich hybridization occurred, the reaction solution maintained its red color. A total of 70 mucous specimens from patients with cervical intraepithelial neoplasia were tested by the AuNP probes sandwich hybridization. The results show that there were 33, 16, 5, and 16 cases detected with HPV-16, HPV-18, both HPV-16 and HPV-18 (HPV-16/HPV-18), and neither HPV-16 nor HPV-18, respectively. In comparison with the specific detection by TaqMan real-time PCR assays for HPV-16, the detection sensitivity and specificity of the AuNP probes sandwich hybridization reached 95% and 90%, respectively, for HPV-16 diagnosis.
SENSING PLATFORM
(2010-12-09)
Lin, Chih-Sheng
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Chuang, Yao-Chen
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Chen, Sz-Hao
A sensing platform includes: a plurality of metal nanoparticles; a plurality of aggregate inducers each comprising first and second functional groups different from each other, and the first functional group of the aggregate inducers being in contact with the metal nanoparticles; and a plurality of recognition molecules for binding the metal nanoparticles and for interacting with a target to recognize the target, wherein the second functional group of the aggregate inducers is free from being in contact with the metal nanoparticles, and is used to induce the metal nanoparticles to aggregate after the recognition molecules interact with the target.
Using oligonucleotide-functionalized Au nanoparticles to rapidly detect foodborne pathogens on a piezoelectric biosensor
(2008-04-01)
Chen, Sz-Hau
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Wu, Vivian C. H.
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Chuang, Yao-Chen
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Lin, Chih-Sheng
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生物科技學系
A circulating-flow piezoelectric biosensor, based on an Au nanoparticle amplification and verification method, was used for real-time detection of a foodbome pathogen, Escherichia coli O157:H7. A synthesized thiolated probe (Probe 1; 30-mer) specific to E. coli O157:H7 eaeA gene was immobilized onto the piezoelectric biosensor surface. Hybridization was induced by exposing the immobilized probe to the E. coli O157:H7 eaeA gene fragment (104-bp) amplified by PCR, resulting in a mass change and a consequent frequency shift of the piezoelectric biosensor. A second thiolated probe (Probe 2), complementary to the target sequence, was conjugated to the An nanoparticles and used as a "mass enhancer" and "sequence verifier" to amplify the frequency change of the piezoelectric biosensor. The PCR products amplified from concentrations of 1.2 x 10(2) CFU/ml of E. coli O157:H7 were detectable by the piezoelectric biosensor. A linear correlation was found when the E. coli O157:H7 detected from 10(2) to 10(6) CFU/ml. The piezoelectric biosensor was able to detect targets from real food samples. (C) 2008 Elsevier B.V. All rights reserved.