Label-free upconversion nanoparticles-based fluorescent probes for sequential sensing of Cu, pyrophosphate and alkaline phosphatase activity.

An efficient near-infrared fluorescence probe has been developed for the sequential detection of Cu, pyrophosphate (PO, PPi), and alkaline phosphatase (ALP), which is based on the "off-on-off" fluorescence switch of branched polyethyleneimine (PEI)-capped NaGdF:Yb/Tm upconversion nanoparticles (UCNPs). The fluorescence is quenched via energy transfer from UCNPs to Cu for the coordination of PEI with Cu. The strong affinity between Cu and PPi leads to the formation of Cu-PPi complex and results in the detachment of Cu from the surface of UCNPs, thus the fluorescence is triggered on. ALP-directed hydrolysis of PPi causes the disassembly of Cu-PPi complex and re-conjugation between Cu with PEI, which leads to the switch-off fluorescence of UCNPs. The system allows sequential analysis of Cu, PPi, and ALP by modulating the switch of the fluorescence of UCNPs with detection limits of 57.8nM, 184nM, and 0.019U/mL for Cu, PPi, and ALP, respectively. By virtue of the NIR feature and excellent biocompatibility, the UCNPs-based probes are suitable for bioimaging. Taking Cu visualization as a model, the nanoprobes have been successfully applied for intracellular imaging of Cu in living cells.