School of Chemistry and Chemical Engineering, Chongqing University of Technology, Chongqing, 400054, PR China.
School of Chemistry and Chemical Engineering, Chongqing University of Technology, Chongqing, 400054, PR China.
Anal Chim Acta. 2021 Mar 1;1148:238195. doi: 10.1016/j.aca.2020.12.070. Epub 2021 Jan 4.
Accurate and sensitive monitoring of the abused antibiotics is vital because excessive antibiotics in human body can cause toxicity to kidney or lead to potential loss of hearing. In this work, we described a label-free and highly sensitive fluorescent aptasensing platform for detecting kanamycin in milk samples based on the synchronization signal amplification of primer exchange reaction (PER) and metal-ion dependent DNAzyme. The target kanamycin binds the aptamer sequence hybridized on a hairpin template and initiates PER for autonomous synthesis of Mg-dependent DNAzyme sequences with aid of Bst-DNA polymerase at isothermal conditions. Such a synthesis process can be repeated many times to produce lots of DNAzymes to cyclically cleave the rA site in the signal hairpin substrates under the assistance of Mg cofactor to liberate numerous free G-quadruplex fragments. The organic dye thioflavin T (ThT) further associates with these G-quadruplex fragments to yield substantially intensified fluorescence for sensitive detection of kanamycin with a low detection limit of 0.36 nM. In addition, the developed aptamer sensing method also shows a good selectivity for kanamycin against other interfering antibiotics, and can realize the monitoring of kanamycin added in milk samples, highlighting its potential for sensitive monitoring of trace amount of kanamycin for food safety applications.
准确且灵敏地监测被滥用的抗生素至关重要，因为人体中过量的抗生素会对肾脏造成毒性，或导致潜在的听力损失。在这项工作中，我们描述了一种基于引物交换反应（PER）和金属离子依赖型 DNA 酶的同步信号放大的无标记、高灵敏荧光适体传感平台，用于检测牛奶样品中的卡那霉素。目标卡那霉素与发夹模板上杂交的适体序列结合，并在等温条件下借助 Bst-DNA 聚合酶启动 PER，从而自主合成 Mg 依赖性 DNA 酶序列。这种合成过程可以重复多次，产生大量的 DNA 酶，在 Mg 辅因子的协助下循环切割信号发夹底物中的 rA 位点，释放出大量游离的 G-四链体片段。有机染料硫黄素 T（ThT）进一步与这些 G-四链体片段结合，产生显著增强的荧光，从而实现对卡那霉素的灵敏检测，检测限低至 0.36 nM。此外，所开发的适体传感方法对卡那霉素具有良好的选择性，可抵抗其他干扰性抗生素，并且可以实现对牛奶样品中添加的卡那霉素的监测，突出了其在食品安全应用中对痕量卡那霉素进行灵敏监测的潜力。
