Colorimetric alkaline phosphatase activity detection by integrating phosphorylation-mediated sulfydryl protection/deprotection and fluorosurfactant stabilized gold nanoparticles

Abstract

Protection/deprotection of thiolates are widely used for the design of sensing systems by producing free sulfydryl (–SH) groups. However, most conventional protection/deprotection strategies reported so far suffer poor protection and/or difficult release of –SH group. In this study, phosphorylation/dephosphorylation strategy that exhibits efficient protection and facile liberation of –SH group via the formation of a phosphate-thiol ester, was utilized to develop colorimetric probe for alkaline phosphatase (ALP) activity evaluation. The proposed probe was composed of a cysteamine S-phosphate as a model thiolate substrate and Zonyl FSN-100 functionalized gold nanoparticles (FSN-AuNPs) as reporters. In the presence of ALP, the cysteamine S-phosphate hydrolyzed to generate cysteamine, which aggregated FSN-AuNPs with detectable spectroscopic changes through nanointerface interactions. The changes in UV–vis absorption spectroscopy allowed a detection limit of ALP at 0.1 unit/L. The specific enzyme catalytic hydrolysis provided high selectivity toward ALP over other proteins, small molecules, and metal ions. The practicability of the sensor was validated by accurate examination of ALP in blood samples. In addition, by integrating the conventional ELISA technique and the proposed phosphorylation tactic, sensitive immunoglobulin G detection was achieved.