Abstract
Albendazole (ABZ) is one of the benzimidazole anthelmintics, and the overuse of ABZ in breeding industry can lead to drug resistance and a variety of toxic effects in humans. Since the residue markers of ABZ are the sum of ABZ and three metabolites (collectively referred to as ABZs), albendazole-sulfone (ABZSO2), albendazole-sulfoxide (ABZSO), and albendazole-2-amino-sulfone (ABZNH2SO2), an antibody able to simultaneously recognize ABZs with high affinity is in urgent need to develop immunoassay for screening purpose. In this work, an unreported hapten, 5-(propylthio)-1H-benzo[d]imidazol-2-amine, was designed and synthesized, which maximally exposed the characteristic sulfanyl group of ABZ to the animal immune system to induce expected antibody. One monoclonal antibody (Mab) that can simultaneously detect ABZs was obtained with IC50 values of 0.20, 0.26, 0.77, and 10.5 μg/L for ABZ, ABZSO2, ABZSO, and ABZNH2SO2 in ic-ELISA under optimized conditions respectively, which has been never achieved in previous reports. For insight into the recognition profiles of the Mab, we used computational chemistry method to parameterize cross-reactive molecules in aspects of conformation, electrostatic fields, and hydrophobicity, revealing that the hydrophobicity and conformation of characteristic group of molecules might be the key factors that together influence antibody recognition with analytes. Furthermore, the practicability of the developed ic-ELISA was verified by detecting ABZs in spiked milk, beef, and liver samples with recoveries of 60% to 108.8% and coefficient of variation (CV) of 1.0% to 15.9%.
Highlights
Albendazole (ABZ, shown in Figure 1a), one of benzimidazoles, is an effective anthelmintic and often used to control soil-transmitted helminth infection in humans and animals
The results shown that the sensitivities of the developed ic-ELISA were times better than those of other reported immunoassays for ABZs (Table 1)
The calculated limit of detection (LOD) of ABZ, ABZSO2, albendazole sulfoxide (ABZSO), and ABZNH2SO2 were at 0.05, 0.05, 0.05, and 0.49 μg/L in milk samples, respectively, and the LODs of ABZNH2SO2 in beef and bovine liver samples were 1.12 and 0.56 μg/kg, which were below the maximum residue limits (MRLs) of ABZs
Summary
Albendazole (ABZ, shown in Figure 1a), one of benzimidazoles, is an effective anthelmintic and often used to control soil-transmitted helminth infection in humans and animals. After administration of ABZ, the kinds of residues that can be monitored depend on the route of administration, target tissue or the detection time after treatment. Residues of ABZ in animal food can lead to embryonic toxicity for consumer, as well as teratogenic and mutagenic effects [9,10]. The European Union (EU) has set the maximum residue limits (MRLs) for ABZs (sum of ABZ, ABZSO, ABZSO2, and ABZNH2SO2, Figure 1a) at. Foods 2021, 10, 3106 limits (MRLs) for ABZs (sum of ABZ, ABZSO, ABZSO2, and ABZNH2SO2, Figure 1a) at 100 μg/kg (L) in milk/muscle/fat, 500 μg/kg in kidney, and 1000 μg/kg in liver [11]. All of the buffers used in the immunoassay have been listed in Supplementary Materials
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