Disposable Amperometric Polymerase Chain Reaction-Free Biosensor for Direct Detection of Adulteration with Horsemeat in Raw Lysates Targeting Mitochondrial DNA.
Anal Chem
; 89(17): 9474-9482, 2017 09 05.
Article
in En
| MEDLINE
| ID: mdl-28746806
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 (â¼16â¯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.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
DNA, Mitochondrial
/
Polymerase Chain Reaction
/
Electrochemical Techniques
/
Meat
Type of study:
Diagnostic_studies
Limits:
Animals
Language:
En
Journal:
Anal Chem
Year:
2017
Document type:
Article
Affiliation country:
Spain
Publication country:
EEUU
/
ESTADOS UNIDOS
/
ESTADOS UNIDOS DA AMERICA
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EUA
/
UNITED STATES
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UNITED STATES OF AMERICA
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US
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USA