Imaging Live Cells with Nonblinking and Nonbleaching NIR-to-Visible Upconverting Nanoparticles

Abstract

Recently, lanthanide ion-doped upconverting nanoparticles (UCNPs) are emerging as promising biological imaging probes thanks to their unique optical properties. The visible luminescence of UCNPs upon the NIR excitation was monitored at the single-particle level using a sensitive wide-field fluorescence microscope system equipped with a 980 nm diode laser and an EMCCD camera. It was found that single UCNPs exhibit neither photoblinking on the millisecond time scale nor photobleaching under long-time continuous illumination. We carried out cellular imaging using biocompatible PEG-lipid coated UCNPs as bioprobes in living HeLa cells. We observed that the UCNPs were internalized and transported to the perinuclear region through endocytic process from the snapshot images taken over several hours. We also found the significant signal loss, presumably due to diffusion and/or exocytosis. Owing to the NIR excitation, the luminescence images were free of autofluorescent background from cellular environment while the cell samples were not significantly damaged. In particular, we could visualize the directed motion of the vesicles, where UCNPs were encapsulated, from single particle tracking in real time, indicating that UCNPs are actively transported by the motor protein, such as dynein or kinesin, moving along the microtubules.