Abstract
A fluorescent immunoassay based on europium nanoparticles (EuNPs-FIA) was developed for the simultaneous detection of antibiotic residues, solving the problems of single target detection and low sensitivity of traditional immunoassay methods. In the EuNPs-FIA, EuNPs were used as indictive probes by binding to anti-tetracyclines monoclonal antibodies (anti-TCs mAb), anti-sulphonamides monoclonal antibodies (anti-SAs mAb) and anti-fluoroquinolones monoclonal antibodies (anti-FQs mAb), respectively. Different artificial antigens were assigned to different regions of the nitrocellulose membrane as capture reagents. The EuNPs-FIA allowed for the simultaneous detection of three classes of antibiotics (tetracyclines, fluoroquinolones and sulphonamides) within 15 min. It enabled both the qualitative determination with the naked eye under UV light and the quantitative detection of target antibiotics by scanning the fluorescence intensity of the detection probes on the corresponding detection lines. For qualitative analysis, the cut-off values for tetracyclines (TCs), fluoroquinolones (FQs) and sulphonamides (SAs) were 3.2 ng/ml, 2.4 ng/ml and 4.0 ng/ml, respectively, which were much lower than the maximum residue limit in food. For quantitative analysis, these ranged from 0.06 to 6.85 ng/ml for TCs, 0.03–5.14 ng/ml for FQs, and 0.04–4.40 ng/ml for SAs. The linear correlation coefficients were higher than 0.97. The mean spiked recoveries ranged from 92.1 to 106.2% with relative standard deviations less than 8.75%. Among them, the three monoclonal antibodies could recognize four types of TCs, seven types of FQs and 13 types of SAs, respectively, and the detection range could cover 24 antibiotic residues with different structural formulations. The results of the detection of antibiotic residues in real samples using this method were highly correlated with those of high performance liquid chromatography (R 2 > 0.98). The accuracy and precision of the EuNPs-FIA also met the requirements for quantitative analysis. These results suggested that this multiplex immunoassay method was a promising method for rapid screening of three families of antibiotic residues.
Highlights
Since the discovery of penicillin in 1929, antibiotics were frequently used as clinical agents to prevent and treat bacterial infections in human and veterinary medicine (Kong et al, 2020)
The detection principle of immunochromatography system was based on the competitive reaction theory (Gong et al, 2014), the principle is based on the competition between target detectors contained in the detection sample and artificial antigen conjugates fixed on nitrocellulose membrane, so as to bind to the monoclonal antibody labeled by EuNPs (Figure 1)
On the contrary, when the sample solution does not contain the target analyte, the monoclonal antibody labeled by EuNPs reacted with artificial antigen on T-line and goat anti-mouse IgG on C-line (Figure 1C)
Summary
Since the discovery of penicillin in 1929, antibiotics were frequently used as clinical agents to prevent and treat bacterial infections in human and veterinary medicine (Kong et al, 2020). The excessive and indiscriminate use of antibiotics in interconnected ecosystems around the world had led to a dramatic rise in drug resistance This concern had been listed by the World Health Organisation as one of the top ten public health threats (Anita et al, 2021). Antibiotics could modulate human mood and increase the risk of depression (mental illness) by altering the gut microbiota and brain-gut axis (Konstantinidis et al, 2020). It may induce a range of genetic effects in humans, such as carcinogenic or allergic reactions (Li et al, 2020)
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