Determination of alkaline phosphatase activity based on enzyme-triggered generation of a thiol and the fluorescence quenching of silver nanoclusters

• Published in: Mikrochimica acta (1966) 2019-03, Vol.186 (3), p.180-180, Article 180
• Main Authors: Luo, Minchuan, Su, Zhu, Wang, Xinyi, Li, Liang, Tu, Yifeng, Yan, Jilin
• Format: Article
• Language: English
• Subjects: Alkaline Phosphatase - blood; Alkaline Phosphatase - chemistry; Amifostine - chemistry; Analytical Chemistry; Characterization and Evaluation of Materials; Chemistry; Chemistry and Materials Science; Enzyme Assays - methods; Enzymes; Fluorescence; Fluorometry - methods; Humans; Limit of Detection; Metal Nanoparticles - chemistry; Metal Nanoparticles - radiation effects; Microengineering; Nanochemistry; Nanotechnology; Original Paper; Phosphatases; Polyethyleneimine - chemistry; Silver - chemistry; Sulfhydryl Compounds - chemical synthesis; Thiols; Ultraviolet Rays
• ISSN: 0026-3672; 1436-5073
• DOI: 10.1007/s00604-019-3301-4

A fluorimetric method is described for the determination of alkaline phosphatase (ALP) activity. It is based on the use of polyethyleneimine-coated silver nanoclusters (AgNCs), which display an intense blue fluorescence peaking at 450 nm (under 375 nm excitation). ALP catalyzes the dephosphorylation of the thiophosphate amifostine to generate a thiol that binds to the AgNCs and causes its fluorescence to be quenched. Under the optimal experimental conditions, fluorescence linearly drops in the 0.08–2.0 U L −1 ALP activity range, and the limit of detection is 0.02 U L −1 . The method was successfully applied to the determination of ALP activity in spiked human serum samples. Graphical abstract Alkaline phosphatase (ALP) catalyzes the degradation of amifostine with a generation a thiol product. The thiol quenches the fluorescence of silver nanoclusters, and a method for the detection of ALP down to 0.02 U L −1 was developed.