Label-free fluorescent detection of alkaline phosphatase with vegetable waste-derived green carbon probes

Abstract

The level of alkaline phosphatase (ALP) is regarded as an important biomarker in medical diagnosis. In this study, a carbon-based fluorescent probes (C-probes) to detect ALP was prepared by modified-ultrasound irradiation from vegetable waste as a starting carbon source. The C-probes are water soluble, non-cytotoxic, biocompatible, and exhibits high fluorescence, which diminishes depending on the amount of ALP. The developed assay relies on Cu2+ induced fluorescence quenching of the C-probes and its inhibition by the very selective coordination of pyrophosphate (PPi) with Cu2+ ions. In the presence of PPi, Cu2+ reacts preferentially with them rather than forming complexes with the C-probes. This phenomenon disappears when PPi undergoes ALP-catalyzed hydrolysis to generate phosphate (Pi), which does not bind to Cu2+ ions. The ALP inhibits the coordination of PPi with Cu2+ and thus causes fluorescence quenching of the C-probes by free Cu2+. Using this strategy, ALP could be analyzed successfully down to 0.25 nM, with a dynamic linear range of 0.5-10 nM. This sensing strategy also revealed a high selectivity for ALP over other protein molecules. Furthermore, the method successfully detected biological ALP in human serum, which verifies the potential of the method for diagnostic and practical purposes. (C) 2018 Elsevier B.V. All rights reserved.