Development of a direct competitive chemiluminescent ELISA for the detection of nitrofurantoin metabolite 1-amino-hydantoin in fish and honey

Analytical Methods Pub Date: 2014-04-10 DOI: 10.1039/C4AY00487F

Abstract

A direct competitive enzyme-linked immunosorbent assay (ELISA) with chemiluminescent (dcCLELISA) detection for 1-amino-hydantoin (AHD) was developed in this study. AHD was derivatised with 4-carboxybenzaldehyde to produce 1-[(4-carbo-benzylidene)-amino]-imidazolidin-2,4-dione (CPAHD). Monoclonal antibodies (MAb) against AHD were prepared through immunization of BALB/c mice with synthesized CPAHD–Jeffamine–BSA as an antigen. Luminol, p-iodophenol, and urea peroxide mixture solution served as the substrate in CLELISA. The specificity of the MAb, estimated as the cross-reactivity values from the dcCLEILISA assay for 1-[(4-nitro-benzylidene)-amino]-imidazolidin-2,4-dione (NPAHD) and CPAHD, was 100% and 39.67%, respectively. All other compounds showed less than 0.01%. The sensitivity of the antibody, estimated as the IC50 value, was 0.60 μg L?1. The limits of detection for dcCLELISA in fish and honey samples were 0.1 and 0.28 μg kg?1, respectively, and the mean recovery values ranged from 83.6% to 94.7% for fortified samples at levels of 0.25–10 μg kg?1 with coefficient of variation values below 15%. Finally, dcCLELISA was compared to a commercial kit in the detection of AHD in spiked fish and honey samples. The immunoassay method described here showed a broad detection range and high sensitivity. It could be used for high-throughput monitoring of AHD in fish and honey samples and possibly other types of food.

Graphical abstract: Development of a direct competitive chemiluminescent ELISA for the detection of nitrofurantoin metabolite 1-amino-hydantoin in fish and honey
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