Organic Solvent-Free Fluorescence Confocal Imaging of Living Cells Using Pure Nanocrystal Forms of Fluorescent Dyes

In this paper we describe an organic solvent-free method for the in vitro fluorescence confocal imaging of living cells using nanocrystals of the hydrophobic fluorescent dyes 3,3'-dioctadecyloxacarbocyanine perchlorate (DiO) and perylene, which we obtained separately using the reprecipitation method. Confocal microscopy imaging revealed that cancer cells took up the dye with high avidity. Under similar treatment conditions, the dyes in their nanocrystal forms or as dimethylsulfoxide solutions provided comparable in vitro fluorescence imaging of cells through confocal microscopy. This nanocrystal formulation approach to in vitro bioimaging in cultures eliminates the need for external solubilizing agents (i.e., organic solvent, surfactant), which usually exhibit cytotoxicity. Our developed technique also allowed the double-staining imaging of cancer cells using DiO and perylene nanocrystals.