1. Homogeneous Quenching Immunoassay for Fumonisin B 1 Based on Gold Nanoparticles and an Epitope-Mimicking Yellow Fluorescent Protein.
- Author
-
Peltomaa R, Amaro-Torres F, Carrasco S, Orellana G, Benito-Peña E, and Moreno-Bondi MC
- Subjects
- Bacterial Proteins analysis, Fluorescence, Luminescent Proteins analysis, Bacterial Proteins chemistry, Epitopes chemistry, Fumonisins analysis, Fumonisins chemistry, Gold chemistry, Immunoassay, Luminescent Proteins chemistry, Metal Nanoparticles chemistry
- Abstract
Homogeneous immunoassays represent an attractive alternative to traditional heterogeneous assays due to their simplicity, sensitivity, and speed. On the basis of a previously identified epitope-mimicking peptide, or mimotope, we developed a homogeneous fluorescence quenching immunoassay based on gold nanoparticles (AuNPs) and a recombinant epitope-mimicking fusion protein for the detection of mycotoxin fumonisin B
1 (FB1 ). The fumonisin mimotope was cloned as a fusion protein with a yellow fluorescent protein that could be used directly as the tracer for FB1 detection without the need of labeling or a secondary antibody. Furthermore, owing to the fluorescence quenching ability of AuNPs, a homogeneous immunoassay could be performed in a single step without washing steps to separate the unbound tracer. The homogeneous quenching assay showed negligible matrix effects in 5% wheat extract and high sensitivity for FB1 detection, with a dynamic range from 7.3 to 22.6 ng mL-1 , a detection limit of 1.1 ng mL-1 , and IC50 value of 12.9 ng mL-1 , which was significantly lower than the IC50 value of the previously reported assay using the synthetic counterpart of the same mimotope in a microarray format. The homogeneous assay was demonstrated to be specific for fumonisins B1 and B2 , as no significant cross-reactivity with other mycotoxins was observed, and acceptable recoveries (86% for FB1 2000 μg kg-1 and 103% for FB1 4000 μg kg-1 ), with relative standard deviation less than 6.5%, were reported from spiked wheat samples, proving that the method could provide a valuable tool for simple analysis of mycotoxin-contaminated food samples.- Published
- 2018
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