Integrated double signal amplification systems with ELISA assay for sensitive detection of tylosin in food

Tylosin (TYL) is a kind of antibiotics which promotes the development of animal husbandry, but it poses potential threats to food safety and human health. Therefore, it is necessary to establish a highly sensitive method for the detection of TYL. Here, an enzyme-linked immunosorbent assay (ELISA) combined with biotin–streptavidin (Bio-SA) system and hybridization chain reaction (HCR) technique (Bio-SA-HCR-ELISA) was developed. In the absence of the TYL, a strong fluorescence was generated through double signal amplification as signal output, while in the presence of TYL, the fluorescence intensity decreased. The results showed that the limit of detection for TYL was 0.39 ng/mL, and this method had a good specificity for 9 common antiboics. Compared to the traditional indirect competitive ELISA, the sensitivity of this method was 6.8 times higher. Additionally, the established double signal amplification Bio-SA-HCR-ELISA was used for detecting TYL in milk and honey, the LOD of this method were 4.9 and 2.5 ng/mL, respectively.

泰乐菌素（Tylosin, TYL）是一类可推动畜牧业发展的抗生素，但同时会对食品安全与人类健康构成潜在威胁。因此，建立高灵敏度的泰乐菌素检测方法十分必要。本研究开发了一种结合生物素-链霉亲和素（biotin–streptavidin, Bio-SA）系统与杂交链反应（hybridization chain reaction, HCR）技术的酶联免疫吸附测定（enzyme-linked immunosorbent assay, ELISA）方法（Bio-SA-HCR-ELISA）。当体系中无泰乐菌素时，该方法通过双重信号放大产生强荧光作为信号输出；当体系中存在泰乐菌素时，荧光强度则会降低。实验结果显示，该方法对泰乐菌素的检出限为0.39 ng/mL，且对9种常见抗生素具备良好的特异性。相较于传统间接竞争ELISA方法，本方法的灵敏度提升了6.8倍。此外，将构建的双重信号放大Bio-SA-HCR-ELISA方法应用于牛奶与蜂蜜样品中的泰乐菌素检测，其在两种基质中的检出限分别为4.9 ng/mL和2.5 ng/mL。