A turn-on fluorescence assay of alkaline phosphatase activity based on an enzyme-triggered conformational switch of G-quadruplex

• Published in: Talanta (Oxford) Vol. 208; p. 120453
• Main Authors: Zhou, Xi, Khusbu, Farjana Yeasmin, Chen, Hanchun, Ma, Changbei
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 01.02.2020
• Subjects: Alkaline phosphatase; Fluorescence assay; G-quadruplex; Thioflavin T; Alkaline phosphatase; Thioflavin T; Fluorescence assay; G-quadruplex
• ISSN: 0039-9140; 1873-3573
• DOI: 10.1016/j.talanta.2019.120453

Herein, a turn-on fluorescence assay was introduced for alkaline phosphatase (ALP) detection based on ThT/G-quadruplex system. The basis of the method is that chelation of guanine bases at the binding sites by Ag+ blocks G-quadruplex formation and decreases the fluorescence intensity sharply. In the presence of ALP, ascorbic acid 2-phosphate (AAP) is hydrolyzed to form ascorbic acid (AA) which in turn reduces Ag+ to Ag0. As a result, the blockage ability of Ag+ is disrupted which augments the fluorescence intensity and relies on the concentration of ALP. Under the optimized parameters (500 nM DNA probe; 6 μM Ag+; 1 mM AAP; 30 min for Ag+ and DNA probe reaction time), fluorescence intensity correlates linear range between 1 and 100 U/L of ALP concentration with the detection limit of 0.503 U/L. In the inhibition assay, 50% of ALP inhibition is caused by Na3VO4 with a concentration of 0.254 mM. Furthermore, the assay was used to detect ALP activity in human serum samples in which the results were significant. Above all, the proposed strategy is potential, facile, and sensitive for analyzing ALP activity and screening ALP inhibitor. A turn-on fluorescence assay has been developed for the detection of alkaline phosphatase based on a conformational switch of a G-quadruplex mediated by an enzyme-triggered silver reduction. [Display omitted] •A label-free fluorescence assay was developed for alkaline phosphatase detection.•It is based on a conformational switch of a G-quadruplex mediated by an enzyme-triggered silver reduction.•This method confers high sensitivity, selectivity and application potential.•It is simple and cost-effective without any labels or complicated operations.