Alkaline phosphatase determination via regulation of enzymatically generated poly(thymine) as a template for fluorescent copper nanoparticle formation
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We propose a new fluorometric method for alkaline phosphatase (ALP) determination. This method is based on the regulation of enzymatically generated poly(thymine) for the preparation of copper nanoparticles (CuNPs). 2′-Deoxythymidine 5′-triphosphate (dTTP) serves as the source for polymerization mediated by terminal deoxynucleotidyl transferase (TdT). This process generates poly(thymine), which acts as the template for synthesis of fluorescent CuNPs. However, if ALP catalyzes the hydrolysis of dTTP, the TdT-mediated polymerization will be disabled. This prevents the formation of CuNPs and causes a drop in fluorescence. The findings were used to design a sensitive and selective fluorometric method for ALP determination. A linear response in the activity range from 0.1 to 20 U L-1 and a limit of quantification of 0.3 U L-1 were obtained. The results indicate that the proposed method can be successfully applied to ALP assay in spiked diluted serum. This demonstrates the method’s reliability and practicability.
KeywordsAlkaline phosphatase Enzymatic polymerization Poly(thymine) Copper nanoparticles 2′-Deoxythymidine 5′-triphosphate
The authors thank the National Natural Science Foundation of China (no. 21405125) and the Natural Science Foundation Project of Chongqing (no. cstc2018jscx-msybX0263) for financial support.
Compliance with ethical standards
All procedures performed in studies involving animals were in accordance with the ethical standards of Chinese laws and guidelines (GKFCZ2001545) and in accordance with the guidelines of the Committee on the Ethics of Animal Experiments of Southwest University [no. (2007)3].
Conflict of interest
The authors declare that they have no competing interests.
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