Biocompatible fluorescent silicon nanocrystals for single-molecule tracking and fluorescence imaging.

ABSTRACT

Fluorescence microscopy is used extensively in cell-biological and biomedical research, but it is often plagued by three major problems with the presently available fluorescent probes photobleaching, blinking, and large size. We have addressed these problems, with special attention to single-molecule imaging, by developing biocompatible, red-emitting silicon nanocrystals (SiNCs) with a 4.1-nm hydrodynamic diameter. Methods for producing SiNCs by simple chemical etching, for hydrophilically coating them, and for conjugating them to biomolecules precisely at a 11 ratio have been developed. Single SiNCs neither blinked nor photobleached during a 300-min overall period observed at video rate. Single receptor molecules in the plasma membrane of living cells (using transferrin receptor) were imaged for ≥10 times longer than with other probes, making it possible for the first time to observe the internalization process of receptor molecules at the single-molecule level. Spatial variations of molecular diffusivity in the scale of 1-2 µm, i.e., a higher level of domain mosaicism in the plasma membrane, were revealed.