Sensitive detection of hexokinase activity by use of Zr(4+)-coated magnetic beads coupled with phenylboronic acid-functionalized upconversion nanophosphors

Hexokinase (HK)-catalyzed glucose phosphorylation plays an important role in the regulation of circulating glucose levels, so aberrant HK activity may result in various human diseases. Herein, we present a new strategy for highly sensitive detection of HK activity by use of Zr(4+)-coated magnetic beads (ZrMBs) for selective capture of HK-induced phospho-glucose coupled with phenylboronic acid-functionalized upconversion nanophosphors (UCNPs) for specific recognition and low-background detection of the phospho-glucoses anchored on the ZrMBs. In this design, ZrMBs exhibit highly selective binding utility for the phospho-glucose, which binds on the surface of ZrMBs while the unphosphorylated glucose does not. In addition, the magnetic nature of ZrMBs allows the simple purification and separation operations. On the other hand, phenylboronic acid-functionalized UCNPs can specifically recognize the phospho-glucoses anchored on the ZrMBs through strong boronic acid-diols interaction, so the UCNPs finally accumulated on the ZrMBs are proportional to the HK activity. The utilization of UCNPs as the signal reporters can greatly reduce the autofluorescence and light scattering interferences owing to their NIR excitation characteristic, which results in a high signal-to-background ratio. Therefore, by combining these distinct advantages of UCNPs and ZrMBs, ultrahigh sensitivity for the detection of HK activity is achieved.