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A fluorometric aptamer method for kanamycin by applying a dual amplification strategy and using double Y-shaped DNA probes on a gold bar and on magnetite nanoparticles

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Abstract

A simple and highly sensitive fluorometric method is described for the determination of the antibiotic kanamycin (Kana) in food. Dual signal amplification is accomplished by making use of double Y-shaped aptamer DNA probes acting as a capture probes and signal amplification probes. The DNA probes were immobilized on a gold bar and on a magnetic bar, respectively. On addition of Kana, the Y-shaped aptamer probe captures Kana and then is disassembled to release two single-stranded DNAs. These trigger target recycling and HCR between the two bars simultaneously. As a result, many long duplex DNA chains are formed in the supernatant. After pulling out the bars and adding the fluorescent intercalating probe SYBR Green I, strong fluorescence (with excitation/emission peaks at 497/525 nm) is induced. The use of such double Y-shaped DNA probes obviously overcomes the unspecific signal amplification by HCR which increases selectivity and sensitivity. This is due to the fact that the hairpin of HCR is separated in being present in different arms of the Y-shaped probe. Under the optimal conditions, the assay has a limit of 0.45 pg·mL−1 for Kana. It was applied to analyze spiked milk, fish and pork samples.

The scheme represents a sensitive fluorometric aptamer-based method to detect kanamycin (Kana). It is making use of a double stirring bar-assisted dual amplification strategy with zero background. Abbreviations: apt: aptamer, AuNPs: gold nanoparticles, HCR: hybridization chain reaction.

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Acknowledgments

This work was supported by National Key R&D Program of China (No.2016YFD0401502), the National Natural Science Foundation of China (No.31871825), the key project of natural science foundation of china (11832013), Natural Science Foundation of Zhejiang (LY19B050001, LY17C200007), Zhejiang Province Welfare Technology Applied Research Project (2017C330040, LGC19B070003, LGN18H300001), Zhejiang provincial Top key discipline of biology (ZS2015005), the Natural Science and Huiming Foundation of Ningbo (2016A610084, 2017A610225, 2017C50035) and K. C. Wong Magna Fund in Ningbo University.

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Authors and Affiliations

  1. Key Laboratory of Animal Protein Food Processing Technology of Zhejiang Province, College of food and medicine, Ningbo University, Ningbo, 315211, China

    Kai Zhang, Jinxuan Cao & Futao Hu

  2. Faculty of material science and chemical engineering, Ningbo University, Ningbo, 315211, China

    Kai Zhang, Yongxiang Wu, Tianhua Li, Ying Wang & Ning Gan

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  1. Kai Zhang
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  2. Jinxuan Cao
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  7. Ning Gan
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Correspondence to Jinxuan Cao.

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Zhang, K., Cao, J., Wu, Y. et al. A fluorometric aptamer method for kanamycin by applying a dual amplification strategy and using double Y-shaped DNA probes on a gold bar and on magnetite nanoparticles. Microchim Acta 186, 120 (2019). https://doi.org/10.1007/s00604-018-3207-6

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