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Journal of Academic Research and Innovation

Abstract

Zearalenone (ZEN), a mycotoxin produced by Fusarium fungi, is a significant food safety concern due to its widespread contamination in cereal crops and its estrogenic effects on both human and animal health. Although traditional analytical methods such as HPLC and ELISA are very sensitive but they are laborious, expensive, and unsuitable for on-site screening. This paper focused on developing a rapid, cost-effective, and fast lateral flow immunoassay (LFIA) for ZEN detection, using gold nanoparticles (AuNPs) as label particles that were immobilized with anti-ZEN antibodies. AuNPs were synthesized using the Turkevich–Frens method, showing a UV–Vis absorption peak at 520 nm and a monodisperse size of 32.00 nm confirmed by DLS. Conjugation conditions were optimized at pH 7.12, 80 mM NaCl for salt stability, and the optimal antibody concentration was determined to be 4 μg/mL. The optimized condition enabled successful passive adsorption of antibodies onto AuNPs, increasing the hydrodynamic diameter to 127.60 nm by DLS, confirming stable AuNP–antibody conjugates for LFIA development. ZEN, being a low molecular weight, was chemically modified and conjugated to BSA via the NHS/EDC chemistry to achieve the ZEN hapten. Successful conjugation of ZEN-BSA was confirmed by UV-Vis spectroscopy with double peaks at 230 nm (ZEN) and 280 nm (BSA), and thin-layer chromatography (TLC) confirmed purity and conjugate separation. Additionally, nuclear magnetic resonance (NMR) spectroscopy gave evidence of the chemical structure of the synthesized hapten, showing the presence of ZEN moieties covalently attached to the carrier protein, confirming its use as a test line antigen. After synthesis and characterization of AuNPs-antibody conjugate and ZEN-BSA hapten, LFIA strips were assembled using a BIODOT system. 1 mg/mL of ZEN-BSA in PBS was coated as test lines, while goat anti-mouse IgG antibody was coated as control lines. Upon drying, LFIA devices were cut into 5 mm strips and evaluated for dipstick performance. When samples were added, AuNP-tagged antibodies migrated through capillary action, producing a visible color signal in the presence of ZEN. The test strips showed strong signal intensity, stability, and adequate sensitivity for rapid, on-site detection of ZEN in food samples. In conclusion, this study provides an optimized and functional LFIA biosensor for ZEN detection by an inexpensive, quick, and visually accessible method. The comprehensive nanoparticle characterization, successful hapten synthesis (validated by TLC and NMR), and proper strip assembly demonstrate the potential of this platform for efficient food toxin monitoring.

Creative Commons License

Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License
This work is licensed under a Creative Commons Attribution-NonCommercial-Share Alike 4.0 International License.

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