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Disposable Amperometric Polymerase Chain Reaction-Free Biosensor for Direct Detection of Adulteration with Horsemeat in Raw Lysates Targeting Mitochondrial DNA

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Ruiz-Valdepeñas Montiel, Victor and Gutiérrez, María L. and Torrente Rodríguez, Rebeca M. and Povedano, Eloy and Vargas, Eva and Reviejo, Julio and Linacero de la Fuente, Rosario and Gallego, Francisco J. and Campuzano, Susana and Pingarrón Carrazón, José Manuel (2017) Disposable Amperometric Polymerase Chain Reaction-Free Biosensor for Direct Detection of Adulteration with Horsemeat in Raw Lysates Targeting Mitochondrial DNA. Analytical Chemistry, 89 (17). pp. 9474-9482. ISSN (Print Edition) 0003-2700 (Web Edition) 1520-6882

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Official URL: http://pubs.acs.org/doi/abs/10.1021/acs.analchem.7b02412



Abstract

A novel electrochemical disposable nucleic acid biosensor for simple, rapid, and specific detection of adulterations with horsemeat is reported in this work. The biosensing platform involves immobilization of a 40-mer RNA probe specific for a characteristic fragment of the mitochondrial DNA D-loop region of horse onto the surface of magnetic microcarriers. In addition, signal amplification was accomplished by using a commercial antibody specific to RNA/ DNA duplexes and a bacterial protein conjugated with a horseradish peroxidase homopolymer (ProtA-HRP40). Amperometric detection at −0.20 V vs Ag pseudoreference electrode was carried out at disposable screen-printed carbon electrodes. The methodology achieved a limit of detection (LOD) of 0.12 pM (3.0 attomoles) for the synthetic target and showed ability to discriminate between raw beef and horsemeat using just 50 ng of total extracted mitochondrial DNA (∼16 660 bp in length) without previous fragmentation. The biosensor also allowed discrimination between 100% raw beef and beef meat samples spiked with only 0.5% (w/w) horse meat (levels established by the European Commission) using raw mitochondrial lysates without DNA extraction or polymerase chain reaction (PCR) amplification in just 75 min. These interesting features made the developed methodology an extremely interesting tool for beef meat screening, and it can be easily adapted to the determination of other meat adulterations by selection of the appropriate specific fragments of the mitochondrial DNA region and capture probes.


Item Type:Article
Subjects:Sciences > Chemistry > Analytic chemistry
ID Code:44693
Deposited On:18 Sep 2017 11:17
Last Modified:05 Aug 2019 08:47

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