Multiplexed modular genetic targeting of quantum dots.
While DNA-directed nanotechnology is now a well-established platform for bioinspired nanoscale assembly in vitro, the direct targeting of various nanomaterials in living biological systems remains a significant challenge. Hybrid biological systems with integrated and targeted nanomaterials may have interesting and exploitable properties, so methods for targeting various nanomaterials to precise biological locations are required. Fluorescence imaging has benefited from the use of nanoparticles with superior optical properties compared to fluorescent organic dyes or fluorescent proteins. While single-particle tracking (SPT) in living cells with genetically encoded proteins is limited to very short trajectories, the high photon output of genetically targeted and multiplexed quantum dots (QDs) would enable long-trajectory analysis of multiple proteins. However, challenges with genetic targeting of QDs limit their application in these experiments. In this report, we establish a modular method for targeting QD nanoparticles selectively to multiple genetically encoded tags by precomplexing QD-streptavidin conjugates with cognate biotinylated hapten molecules. This approach enables labeling and SPT of multiple genetically encoded proteins on living cells at high speed and can label expressed proteins in the cytosol upon microinjection into living cells. While we demonstrate labeling with three distinct QD conjugates, the approach can be extended to other specific hapten-affinity molecule interactions and alternative nanoparticles, enabling precise directed targeting of nanoparticles in living biological systems.
Department of Chemistry, ‡Molecular Biosensor and Imaging Center, §Department of Physics, ∥Lane Center for Computational Biology, and #Department of Biological Sciences, Carnegie Mellon University , Pittsburgh, Pennsylvania, United States.
Saumya
S
Saurabh
Lauren E
LE
Beck
Suvrajit
S
Maji
Catherine J
CJ
Baty
Yi
Y
Wang
Qi
Q
Yan
Simon C
SC
Watkins
Marcel P
MP
Bruchez
D002460:Cell Line; D004247:DNA / Q000737:chemistry*; D036103:Nanotechnology; D011506:Proteins / Q000737:chemistry*; D045663:Quantum Dots
D011506:Proteins; D004247:DNA
genetic targeting; membrane protein; quantum dots; single-molecule imaging