Gang Ruan, Chemical and Biomolecular Engineering, the Ohio State University, 140 W 19th Ave, Columbus, OH 43210, Amit Agrawal, Health Sciences and Technology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139, and Shuming Nie, Biomedical Enigneering, Georgia Institute of Technology and Emory University, 1639 Pierce Drive Suite 2001, Atlanta, GA 30322.
Single molecule studies can lead to insights not available from examining bulk behaviors due to ensemble averaging effects. Although imaging of single molecules in vitro and in live cell membrane by fluorescence microscopy has been extensively reported, single molecule imaging in live cell cytoplasm remains technically challenging. Single quantum dots have similar sizes as those of biomolecules and have many desirable fluorescent properties, thus offering possibilities for single molecule imaging (if quantum dots are linked with biomolecules). To the best of our knowledge, imaging of single quantum dots in the cytoplasm of live cells has not been reported prior to the present work, due to a number of challenges such as cellular delivery and nanoparticle aggregation.
We have obtained movies of single quantum dots in live cell cytoplasm by combining toxin delivery technique, optimized nanoparticle coating and dynamic confocal imaging. The status of single quantum dots is confirmed by the fluorescence intermittency of these nanoparticles. Interestingly, the aggregation of quantum dots in cytoplasm is found to be dramatically more severe than that in an extracellular environment. Aggregation of nanoparticles in the cytoplasm is greatly reduced by adjusting the surface chemistry of nanoparticles. In addition, the cell type used is also found to affect the nanoparticle aggregation behavior. We have further used thus-obtained single quantum dots to track the dynamics of motor proteins in live cells.