Colorimetric oligonucleotide array for genotyping of hepatitis C virus based on the 5′ non-coding region

Mao, Hongju; Lu, Zhongmin; Zhang, Honglian; Liu, Kangdong; Zhao, Jianlong; Jin, Gen-Di; Gu, Shaohua; Yang, Mengsu

doi:10.1016/j.cca.2007.09.009

Cited by 9 publications

(6 citation statements)

References 40 publications

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“…Although these results are encouraging, the performance of the DNA/DNA hybridization sensor or genosensor at detecting targets in a real-life and complex biological sample, such as a PCR product which has not been purified, may be entirely different. Another point concerns the dilution at which the amplicons are tested in the majority of studies: amplicons are usually tested at low dilutions from 1/3 to 1/40, ,,,− compared to our study in which, despite the amplicons being diluted by a factor of 100, a strong signal was observed.…”

Section: Resultsmentioning

confidence: 99%

“…ELOSA has already been applied to HCV genotyping in the 5′-NCR region . Moreover, two oligonucleotide arrays based on the DNA/DNA hybridization principle were reported on HCV genotyping in the 5′-NCR and NS5b regions. In these studies, the amplicons were produced using semi-nested or nested PCR , and were used at dilutions of 1/3, 1/10, and 1/50 .…”

Section: Resultsmentioning

confidence: 99%

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Development of Innovative and Versatile Polythiol Probes for Use on ELOSA or Electrochemical Biosensors: Application in Hepatitis C Virus Genotyping

et al. 2013

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The aim of this study was to develop versatile diagnostic tools based on the use of innovative polythiolated probes for the detection of multiple viruses. This approach is compatible with optical enzyme-linked oligosorbent assay (ELOSA) or electrochemical (biosensors) detection methods. The application targeted here concerns the rapid genotyping of Hepatitis C virus (HCV). HCV genotyping is one of the predictive parameters currently used to define the antiviral treatment strategy and is based on the sequencing of the viral NS5b region. Generic and specific NS5b amplicons were produced by real-time polymease chain reaction (RT-PCR) on HCV(+) human plasma. Original NS5b probes were designed for genotypes 1a/1b, 2a/2b/2c, 3a, and 4a/4d. Robust polythiolated probes were anchored with good efficacy on maleimide-activated microplates (MAM) and gold electrodes. Their grafting on MAM greatly increased the sensitivity of the ELOSA test which was able to detect HCV amplicons with good sensitivity (10 nM) and specificity. Moreover, the direct and real-time electrochemical detection by differential pulse voltammetry enabled a detection limit of 10 fM to be reached with good reproducibility. These innovative polythiolated probes have allowed us to envisage developing flexible, highly sensitive, and easy-to-handle platforms dedicated to the rapid screening and genotyping of a wide range of viral agents.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Development of Innovative and Versatile Polythiol Probes for Use on ELOSA or Electrochemical Biosensors: Application in Hepatitis C Virus Genotyping

et al. 2013

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“…However, these screening approaches have their distinct limitations: detection of anti‐HCV antibodies such as by the colloidal gold labeled method has modest ability to diagnose early HCV infection due to delayed emergence of the specific antibodies in blood. On the other hand, although HCV‐RNA detection techniques such as real‐time PCR and sequencing of viral genome are the most reliable diagnostic tools for HCV infection, they require expensive laboratory facilities, well‐trained personnel, and are time consuming. Therefore, these shortcomings limit the promotion and application of these techniques for early HCV diagnosis and treatment, especially in resource‐limited areas.…”

Section: Introductionmentioning

confidence: 99%

“…On the other hand, although HCV-RNA detection techniques such as real-time PCR and sequencing of viral genome [24][25][26][27] are the most reliable diagnostic tools for HCV infection, they require expensive laboratory facilities, well-trained personnel, and are time consuming.…”

mentioning

confidence: 99%

Detection of HCV genotypes 1b and 2a by a reverse transcription loop‐mediated isothermal amplification assay

Zhao

Liu

Sun

2017

Journal of Medical Virology

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Hepatitis C virus (HCV) genotypes 1b and 2a are the major cause of liver disease in northern China; however, conventional detection tools are labor-consuming, technically demanding, and costly. Here, we assessed the specificity, sensitivity, and clinical utility of reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detection of HCV genotypes 1b and 2a. Firstly, clinical samples were collected from HCV genotype 1b and 2a infected patients and the RNA were extracted. Secondly, specificity of RT-LAMP assay for detection HCV genotypes 1b and 2a were tested against viral genomes of other hepatitis viruses. Sensitivity of RT-LAMP assay was determined using serial dilutions of standard HCV genotypes 1b and 2a. The amplified products were detected by both electrophoresis and calcein/Mn -dependent visual methods. Finally, we compared the clinical detection rate of RT-LAMP to that of real-time PCR. RT-LAMP assay showed high specificity to detect HCV genotypes 1b and 2b since there was no cross-reactivity with other hepatitis viruses. Sensitivity of RT-LAMP was 100 IU/mL for both genotypes detected by either electrophoresis or calcein/Mn -dependent visual methods. The detection rate of RT-LAMP assay in clinical samples was also comparable to that of real-time PCR without significant difference between the both assays. This study proposes a newly developed RT-LAMP assay for detection of HCV genotypes 1b and 2a. RT-LAMP is highly specific, sensitive, and simple diagnostic tool which would be useful for screening and early diagnosis of HCV especially in resource-limited environments.

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“…Oligonucleotide arrays have emerged as a powerful and convenient tool for parallel and high-throughput experimentation in molecular biological research − . Array technology has considerably simplified the process involved in disease diagnosis and gene mapping, as it offers advantages of using a very small amount of biological probes (nanomoles and picomoles), along with the capability of analyzing thousands of analytes in parallel .…”

Section: Introductionmentioning

confidence: 99%

A Facile Method for the Construction of Oligonucleotide Microarrays

et al. 2008

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In recent years, the oligonucleotide-based microarray technique has emerged as a powerful and promising tool for various molecular biological studies. Here, a facile protocol for the construction of an oligonucleotide microarray is demonstrated that involves immobilization of oligonucleotide-trimethoxysilyl conjugates onto virgin glass microslides. The projected immobilization strategy reflects high immobilization efficiency ( approximately 36-40%) and signal-to-noise ratio ( approximately 98), and hybridization efficiency ( approximately 32-35%). Using the proposed protocol, aminoalkyl, mercaptoalkyl, and phosphorylated oligonucleotides were immobilized onto virgin glass microslides. Briefly, modified oligonucleotides were reacted first with 3-glycidyloxypropyltriethoxysilane (GOPTS), and subsequently, the resultant conjugates were directly immobilized onto the virgin glass surface by making use of silanization chemistry. The constructed microarrays were then used for discrimination of base mismatches. On subjecting to different pH and thermal conditions, the microarray showed sufficient stability. Application of this chemistry to manufacture oligonucleotide probe-based microarrays for detection of bacterial meningitis is demonstrated. Single-step reaction for the formation of conjugates with the commercially available reagent (GOPTS), omission of capping step and surface modification, and efficient immobilization of oligonucleotides onto the virgin glass surface are the key features of the proposed strategy.

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Colorimetric oligonucleotide array for genotyping of hepatitis C virus based on the 5′ non-coding region

Cited by 9 publications

References 40 publications

Development of Innovative and Versatile Polythiol Probes for Use on ELOSA or Electrochemical Biosensors: Application in Hepatitis C Virus Genotyping

Development of Innovative and Versatile Polythiol Probes for Use on ELOSA or Electrochemical Biosensors: Application in Hepatitis C Virus Genotyping

Detection of HCV genotypes 1b and 2a by a reverse transcription loop‐mediated isothermal amplification assay

A Facile Method for the Construction of Oligonucleotide Microarrays

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