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Fluorescent assay of alkaline phosphatase activity via atom transfer radical polymerization

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Abstract

A novel fluorescence assay is proposed through activators regenerated by electron transfer atom transfer radical polymerization (ARGET ATRP) strategy for alkaline phosphatase (ALP) activity detection. First of all, 2-bromo-2-methylpropionic acid (BMP) was employed as the initiator to modify on the surface of the magnetic nanoparticle (Fe3O4-MNP) by amide bonding. Then, ascorbic acid (AA) produced by ALP catalyzed the phosphate group removal from L-ascorbic acid 2-phosphate sesquimagnesium salt hydrate (AAPS), which underwent a redox reaction with Cu(II) and the product Cu(I) triggered the ARGET ATRP reaction. Finally, a strong fluorescent signal could be detected at 514 nm due to numerous fluorescent monomers being grafted to the Fe3O4-MNPs surface (Ex = 490 nm, Em = 514 nm). Under optimal experimental conditions, the linear range of this fluorometric assay for ALP activity was 1–80 mU mL−1, and the detection limit was 0.68 mU mL−1. The method exhibited excellent selectivity and satisfactory results were obtained in the inhibition rate and human serum experiments. Therefore, this ALP activity detection strategy has great potential for clinically relevant disease detection and drug screening.

Graphical abstract

A novel fluorescence strategy for alkaline phosphatase activity detection based on the dephosphorylation property of alkaline phosphatase and ARGET ATRP reaction.

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Acknowledgements

This work was supported by the project of tackling of key scientific and technical problems in Henan Province (202102310149).

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Authors and Affiliations

  1. Pharmacy College, Henan University of Chinese Medicine, Zhengzhou, 450046, People’s Republic of China

    Jing Lu, Daoxiang Li, Lele Ma, Mingsan Miao & Yanju Liu

  2. School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing, 210094, People’s Republic of China

    Jinming Kong

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  1. Jing Lu
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  2. Daoxiang Li
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  3. Lele Ma
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  4. Mingsan Miao
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Correspondence to Mingsan Miao, Yanju Liu or Jinming Kong.

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Lu, J., Li, D., Ma, L. et al. Fluorescent assay of alkaline phosphatase activity via atom transfer radical polymerization. Microchim Acta 189, 84 (2022). https://doi.org/10.1007/s00604-022-05189-6

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