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

摘要

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.