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Organic and Biologically Conjugated Luminescent Quantum Dots


We have designed highly luminescent surface-functionalized CdSe-ZnS semiconductor nanocrystals (quantum dots, QDs) that are stable in a variety of solvents and compatible with biological manipulations (Figure 1). We have also developed a robust non-covalent self-assembly strategy to couple these QDs to a range of either genetically engineered or naturally occurring bio-molecules.1,2 The self-assembly strategy was employed in conjunction with molecular adaptor proteins (bridging proteins) to form stable QD-antibody conjugates and we successfully used them in developing fluoro-immunoassays for the detection of various analytes including small molecule explosives in realistic conditions (Figure 2).2 Multiplexed immunoassays employing our designed QD-antibody conjugates were developed to simultaneously detect up to four relevant soluble analytes: cholera toxin, ricin, shiga like toxin 1, and staphylococcal enterotoxin B.

The highly luminescent water-soluble QDs and QD-bioconjugates developed at NRL have been employed as powerful investigative tools to probe real-time long-term processes in live cells. We have shown that QDs can be easily ingested by live cells through the endocytotic pathway, with no deleterious effects on the cell functions and/or development. Cultures of live cells labeled with QDs using this technique and fed nutrients remained fluorescent during a full cycle of growth and division (about a week). Slime mold cells, which are known to form large scale aggregates of cells (slug-like creatures) when starved, behaved normally even when full of our glowing surface functionalized QDs, indicating that these inorganic fluorophores did not interfere with the cells' ability to develop or to interact with each other (Figure 3a).3 In addition, we have used our QD-antibody conjugate strategy to specifically label mammalian cells. In one example, when QD-antibody conjugates were added to a culture containing cells that are highly enriched with a target receptor, the antibody labeled only its target on the surface of cells that expressed that target protein, demonstrating the quantum dots' specificity (Figure 3b).3 This aspect of our research effort was carried out in collaboration with the group of professor Sanford Simon at the Rockefeller University.

Recently we have taken advantage of some of the unique properties of these inorganic fluorophores (namely strong resistance to degradation and the flexibility of exciting them anywhere below the emission band) to develop fluorescence assays based on Förster Resonance Energy Transfer (FRET). Due to their rather large size (comparable to that of a protein), QD-bioreceptor conjugates offer a unique configuration where a single QD donor can interact simultaneously with several acceptors arrayed quasi-symmetrically around the QD center, resulting in a very substantial improvement in FRET efficiency even when using acceptors that have modest spectral overlap with the QD. This represents a unique advantage in our design for QD-based FRET assays. We used these findings to develop the first prototype of an efficient nanoscale biosensor based on FRET between a QD donor and an array of dyelabeled protein acceptors (Figure 4).4 Sensors of this type promise to be sensitive and regenerable. Future work will further exploit the novel photophysics of QDs to expand the repertoire of their uses. One exciting example is the construction of nanoscale sensors based on the FRET between surfaceimmobilized QD donors conjugated to engineered anti-TNT antibody fragments that are labeled with TNT analogs. These sensors can be integrated into devices for the continuous monitoring of TNT in seawater.

  1. J. Am. Chem. Soc. 122, 12142-12150 (2000).
  2. Analytical Chemistry 74, 841-847 (2002).
  3. Nature Biotechnology 21, 47-51 (2003).
  4. Nature Materials 2, 630-638 (2003).

Contact the Principal Investigator, Hedi Mattoussi, for more information

 

 
   
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