1. Development of innovative and versatile polythiol probes for use on ELOSA or electrochemical biosensors: application in hepatitis C virus genotyping
- Author
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Myriam Lereau, Julie Mayen, Carole Chaix, Jean-Jacques Vasseur, Chantal Fournier-Wirth, Carole Farre, Albert Meyer, Jean-François Cantaloube, François Morvan, Vincent Dugas, Laboratoire TransDiag-Sécurité Transfusionnelle et Innovation Diagnostique, Etablissement Français du Sang Pyrénées-Méditerranée, Institut des Biomolécules Max Mousseron [Pôle Chimie Balard] (IBMM), Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université de Montpellier (UM)-Ecole Nationale Supérieure de Chimie de Montpellier (ENSCM), Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), TECHSEP - TECHniques de SEParations (2011-2014), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Etablissement Français du Sang - Alpes-Méditerranée (EFS - Alpes-Méditerranée), Etablissement Français du Sang, SIMS - Surfaces-(bio)Interfaces - Micro & Nano Systèmes (2011-2014), ANR-09-PIRI-0023 VirProbe, LyonBio-Pole, Eurobiomed, and ANR-09-PIRI-0023,VirProbe,Conception d'un microsystème basé sur des sondes électrochimiques ultra-sensibles pour une multi-détection sans marquage d'acides nucléiques: application au génotypage HCV(2009)
- Subjects
Genotype ,Polymers ,Hepatitis C virus ,RT-PCR ,Computational biology ,Biosensing Techniques ,Hepacivirus ,010402 general chemistry ,medicine.disease_cause ,Real-Time Polymerase Chain Reaction ,01 natural sciences ,Analytical Chemistry ,03 medical and health sciences ,chemistry.chemical_compound ,optical enzyme-linked oligosorbent assay ,medicine ,Electrochemical biosensor ,Humans ,Sulfhydryl Compounds ,Genotyping ,NS5B ,030304 developmental biology ,Detection limit ,0303 health sciences ,Molecular Structure ,Chemistry ,[CHIM.ORGA]Chemical Sciences/Organic chemistry ,real-time polymease chain reaction ,Electrochemical Techniques ,Amplicon ,Molecular biology ,NS5b ,0104 chemical sciences ,ELOSA ,HCV ,Differential pulse voltammetry ,Oligonucleotide Probes ,Biosensor - Abstract
International audience; 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.
- Published
- 2013
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