Sensitive Fluorescence Assay for the Detection of Alkaline Phosphatase Based on a Cu2+-Thiamine System

Abstract

The authors describe a novel, facile, and sensitive fluorometric strategy based on a Cu2+-thiamine (Cu2+-TH) system for the detection of alkaline phosphatase (ALP) activity and inhibition. The principle of the method is as follows. Under a basic conditions, TH, which does not exhibit a fluorescence signal, is oxidized into fluorescent thiochrome (TC) by Cu2+. Ascorbic acid 2-phosphate (AAP), which is the enzyme substrate, is hydrolyzed to produce ascorbic acid (AA) by ALP. The newly formed AA then reduces Cu2+ to Cu+, which prevents the oxidation of TH by Cu2+; as a result, the fluorescent signal becomes weaker. On the contrary, in the absence of ALP, AAP cannot reduce Cu2+; additions of Cu2+ and TH result in a dramatic increase of the fluorescent signal. The sensing strategy displays brilliant sensitivity with a detection limit of 0.08 U/L, and the detection is linear in the concentration range of 0.1 to 100 U/L. This approach was successfully applied to ALP activity in human serum samples, indicating that it is reliable and may be applied to the clinical diagnosis of ALP-related diseases.

Alkaline Phosphatase Electrochemical Micro-Sensor Based on 3D Graphene Networks for the Monitoring of Osteoblast Activity

“…To date, various detection strategies have been developed for measuring ALP activity, such as colorimetry [ 7 , 8 ], surface-enhanced resonance Raman scattering [ 9 , 10 ], and fluorescence assay [ 11 , 12 , 13 ]. Despite their reliability and sensitivity, many of the testing methods are time-consuming, requiring professional operators and sophisticated instruments.…”

NPA-Cu2+ Complex as a Fluorescent Sensing Platform for the Selective and Sensitive Detection of Glyphosate

“…Fluorescence sensors such as nanoclusters [ 19 , 20 , 21 ], quantum dots [ 22 , 23 , 24 ], and metal-organic frameworks have been used to detect glyphosate [ 25 ]. Currently, copper ion complexes are widely used due to their novelty, sensitivity, low toxicity, and membrane permeability [ 26 , 27 , 28 ]. A coumarin derivative and Cu 2+ complex system was developed that selectively detected glyphosate with a good linear relationship of 0.02–1.50 μg/mL [ 29 ].…”

An Exonuclease I-Aided Turn-Off Fluorescent Strategy for Alkaline Phosphatase Assay Based on Terminal Protection and Copper Nanoparticles

“…Studies have shown that ALP is able to catalyze alkaline hydrolysis of nucleic acids, proteins, and some small molecules, which are phosphate [ 1 , 3 , 4 , 5 ]. Owing to the indispensable role in many physiological processes such as cell cycle, growth, apoptosis, and signal transduction, ALP is closely connected to multiple human diseases, especially bone and hepatic diseases [ 5 , 6 , 7 ]. The concentration of ALP in healthy people’s serum is 46–190 U/L [ 8 ].…”