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A Ratiometric Fluorescence Biosensor for Detection of Alkaline Phosphatase Via an Advanced Chemometric Model

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Abstract

In this paper, a ratiometric fluorescence biosensor was introduced for alkaline phosphatase (ALP) detection based on 2-aminopurine (2-Amp) and thioflavin T (ThT)-G-quadruplex system. We designed a special DNA (5'-AGGGTTAGGGTTAGGGTTAGGGAAA/i2-Amp/AAAA-PO4-3', AP) modified with a phosphate moiety at the 3’-end, G-quadruplex at the 5’-end, and a fluorophore (2-Amp) in the middle. In the absence of ALP, the G-rich AP strand could be prone to fold into G-quadruplex structures in the presence of K+. Then, ThT combined with G-quandruplex, resulting in the enhancement of fluorescence emission peak at 485 nm. However, ALP-mediated hydrolysis of the 3’-phosphoryl end promoted the cleavage of AP by the exonuclease I (Exo I), releasing 2-Amp which displayed a strong fluorescence emission peak at 365 nm. Moreover, the quantitative fluorescence model (QFM) was derived for the analysis of the fluorescence measurements obtained by the proposed ratiometric fluorescent biosensor. With the aid of the advanced model, the proposed ratiometric fluorescent biosensor possessed satisfactory results for the detection of ALP in the human serum samples, with accuracy comparable to that of the reference method—the commercial ALP assay kit. Under the optimized experimental conditions, this method exhibited good selectivity and higher sensitivity, and the detection limit was found to be as low as 0.017 U/L. Therefore, it is reasonable to expect that the method had a great potential to detect ALP quantitatively in clinical diagnosis.

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Data Availability

All data generated or analyzed during this study are included in this published article (and its supplementary information file).

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Funding

Open access funding is provided by the Hunan Provincial Education Department Scientific Research Outstanding Youth Project (No. 22B0579), the National Natural Science Foundation of China (No. 22204049), the Natural Science Foundation of Hunan Province (No. 2022JJ40042) and the Natural Science Foundation of Changsha (No. kq2202143).

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Authors and Affiliations

  1. Hunan Key Lab of Biomedical Materials and Devices, College of Life Sciences and Chemistry, Hunan University of Technology, Zhuzhou, 412007, PR China

    Yao Chen, Jing-Jing Han, Bo-Wen Li, Li-Bo Nie & Ying Tang

  2. State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082, China

    Yao Chen, Ying Tang & Tong Wang

Authors
  1. Yao Chen
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Contributions

Yao Chen: conceptualization, methodology, data curation, software, writing-original draft. Jing-Jing Han: methodology, data curation. Bo-Wen Li: validation. Li-Bo Nie: formal analysis, writing-review & editing. Ying Tang: conceptualization, investigation. Tong Wang: funding acquisition, supervision.

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Correspondence to Ying Tang or Tong Wang.

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Chen, Y., Han, JJ., Li, BW. et al. A Ratiometric Fluorescence Biosensor for Detection of Alkaline Phosphatase Via an Advanced Chemometric Model. J Fluoresc (2023). https://doi.org/10.1007/s10895-023-03445-3

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