Multiresidue Detection of Fluoroquinolones: Specificity Engineering of a Recombinant Antibody with Oligonucleotide-Directed Mutagenesis

Screening of a group of antibiotics from foodstuffs has traditionally relied on sophisticated chemical or physical analysis methods, such as liquid chromatography and mass spectrometric applications. The equipment for these techniques is expensive and not always applicable for high throughput screen...

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Bibliographic Details
Published in:Journal of agricultural and food chemistry 2013-12, Vol.61 (49), p.11981-11985
Main Authors: Leivo, Janne, Lamminmäki, Urpo, Lövgren, Timo, Vehniäinen, Markus
Format: Article
Language:English
Subjects:
Animals
Anti-Bacterial Agents - analysis
Antibodies - analysis
Antibodies - genetics
Antibodies - metabolism
Biological and medical sciences
Drug Residues - analysis
Fluoroquinolones - analysis
Food Contamination - analysis
Fundamental and applied biological sciences. Psychology
Immunoassay - instrumentation
Immunoassay - methods
Milk - chemistry
Mutagenesis, Site-Directed
Oligonucleotides - genetics
Protein Engineering
Recombinant Proteins - analysis
Recombinant Proteins - genetics
Recombinant Proteins - metabolism
Species Specificity
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Summary:Screening of a group of antibiotics from foodstuffs has traditionally relied on sophisticated chemical or physical analysis methods, such as liquid chromatography and mass spectrometric applications. The equipment for these techniques is expensive and not always applicable for high throughput screening. There is a need for an easy and cost efficient detection method for simultaneous screening of structurally similar compounds. Here we describe the engineering of a recombinant antibody which was subjected to oligonucleotide targeted random mutagenesis to emphasize the generic specificity of fluoroquinolone binding. Phage display together with small sized fluoroquinolone derivatives was used to find antibodies of high affinity and generic specificity. The most improved antibody was used to develop a time-resolved fluorescence immunoassay which was further optimized and applied for the detection of fluoroquinolone residues from spiked whole milk samples. The assay can be used to efficiently screen all European Agency for the Evaluation of Medicinal Products (EMEA) controlled fluoroquinolones from whole milk samples with detection levels ranging from 0.2 to 68 μg L–1.
ISSN:0021-8561
1520-5118
DOI:10.1021/jf403715n