Quantum dot-loaded monofunctionalized DNA icosahedra for single-particle tracking of endocytic pathways


Functionalization of quantum dots (QDs) with a single biomolecular tag using traditional approaches in bulk solution has met with limited success. DNA polyhedra consist of an internal void bounded by a well-defined three-dimensional structured surface. The void can house cargo and the surface can be functionalized with stoichiometric and spatial precision. Here, we show that monofunctionalized QDs can be realized by encapsulating QDs inside DNA icosahedra and functionalizing the DNA shell with an endocytic ligand. We deployed the DNA-encapsulated QDs for real-time imaging of three different endocytic ligands—folic acid, galectin-3 (Gal3) and the Shiga toxin B-subunit (STxB). Single-particle tracking of Gal3- or STxB-functionalized QD-loaded DNA icosahedra allows us to monitor compartmental dynamics along endocytic pathways. These DNA-encapsulated QDs, which bear a unique stoichiometry of endocytic ligands, represent a new class of molecular probes for quantitative imaging of endocytic receptor dynamics.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Encapsulation of QDs within DNA icosahedra.
Figure 2: Cellular validation of atomistic model of the DNA icosahedron.
Figure 3: Binding of IQDGal3 to the plasma membranes of cells.
Figure 4: TEM studies reveal that IQDGal3 is endocytosed through CLICs.
Figure 5: Single-particle tracking of IQDSTxB in live cells.


  1. 1

    Kairdolf, B. A. et al. Semiconductor quantum dots for bioimaging and biodiagnostic applications. Annu. Rev. Anal. Chem. 6, 143–162 (2013).

    CAS  Article  Google Scholar 

  2. 2

    You, C. et al. Self-controlled monofunctionalization of quantum dots for multiplexed protein tracking in live cells. Angew. Chem. Int. Ed. 49, 4108–4112 (2010).

    CAS  Article  Google Scholar 

  3. 3

    Clarke, S. et al. Covalent monofunctionalization of peptide-coated quantum dots for single-molecule assays. Nano Lett. 10, 2147–2154 (2010).

    CAS  Article  Google Scholar 

  4. 4

    You, C. et al. Electrostatically controlled quantum dot monofunctionalization for interrogating the dynamics of protein complexes in living cells. ACS Chem. Biol. 8, 320–326 (2013).

    CAS  Article  Google Scholar 

  5. 5

    Carstairs, H. M. J., Lymperopoulos, K., Kapanidis, A. N., Bath, J. & Turberfield, A. J. DNA monofunctionalization of quantum dots. ChemBioChem 10, 1781–1783 (2009).

    CAS  Article  Google Scholar 

  6. 6

    Howarth, M. et al. Monovalent, reduced-size quantum dots for imaging receptors on living cells. Nat. Methods 5, 397–399 (2008).

    CAS  Article  Google Scholar 

  7. 7

    Farlow, J. et al. Formation of targeted monovalent quantum dots by steric exclusion. Nat. Methods 10, 1203–1205 (2013).

    CAS  Article  Google Scholar 

  8. 8

    Bhatia, D. et al. Icosahedral DNA nanocapsules by modular assembly. Angew. Chem. Int. Ed. 48, 4134–4137 (2009).

    CAS  Article  Google Scholar 

  9. 9

    Pan, K. et al. Lattice-free prediction of three-dimensional structure of programmed DNA assemblies. Nat. Commun. 5, 5578 (2014).

    CAS  Article  Google Scholar 

  10. 10

    Zhang, C. et al. Exterior modification of a DNA tetrahedron. Chem. Commun. 46, 6792–6794 (2010).

    CAS  Article  Google Scholar 

  11. 11

    Angell, C., Xie, S., Zhang, L. & Chen, Y. DNA nanotechnology for precise control over drug delivery and gene therapy. Small http://dx.doi.org/10.1002/smll.201502167 (2016).

  12. 12

    Bhatia, D., Surana, S., Chakraborty, S., Koushika, S. P. & Krishnan, Y. A synthetic icosahedral DNA-based host-cargo complex for functional in vivo imaging. Nat. Commun. 2, 339 (2011).

    Article  Google Scholar 

  13. 13

    Surana, S., Bhatia, D. & Krishnan, Y. A method to study in vivo stability of DNA nanostructures. Methods 64, 94–100 (2013).

    CAS  Article  Google Scholar 

  14. 14

    Edwardson, T. G. W., Carneiro, K. M. M., McLaughlin, C. K., Serpell, C. J. & Sleiman, H. F. Site-specific positioning of dendritic alkyl chains on DNA cages enables their geometry-dependent self-assembly. Nat. Chem. 5, 868–875 (2013).

    CAS  Article  Google Scholar 

  15. 15

    McLaughlin, C. K. et al. Three-dimensional organization of block copolymers on ‘DNA-minimal’ scaffolds. J. Am. Chem. Soc. 134, 4280–4286 (2012).

    CAS  Article  Google Scholar 

  16. 16

    Perrault, S. D. & Shih, W. M. Virus-inspired membrane encapsulation of DNA nanostructures to achieve in vivo stability. ACS Nano 8, 5132–5140 (2014).

    CAS  Article  Google Scholar 

  17. 17

    Lee, H. et al. Molecularly self-assembled nucleic acid nanoparticles for targeted in vivo siRNA delivery. Nat. Nanotech. 7, 389–393 (2012).

    CAS  Article  Google Scholar 

  18. 18

    Jaiswal, J. K. & Simon, S. M. Imaging single events at the cell membrane. Nat. Chem. Biol. 3, 92–98 (2007).

    CAS  Article  Google Scholar 

  19. 19

    Alivisatos, A. P., Gu, W. & Larabell, C. Quantum dots as cellular probes. Annu. Rev. Biomed. Eng. 7, 55–76 (2005).

    CAS  Article  Google Scholar 

  20. 20

    Johannes, L., Parton, R. G., Bassereau, P. & Mayor, S. Building endocytic pits without clathrin. Nat. Rev. Mol. Cell Biol. 42, 1–11 (2015).

    Google Scholar 

  21. 21

    Carion, O., Mahler, B., Pons, T. & Dubertret, B. Synthesis, encapsulation, purification and coupling of single quantum dots in phospholipid micelles for their use in cellular and in vivo imaging. Nat. Protoc. 2, 2383–2390 (2007).

    CAS  Article  Google Scholar 

  22. 22

    Mahler, B. et al. Towards non-blinking colloidal quantum dots. Nat. Mater. 7, 659–664 (2008).

    CAS  Article  Google Scholar 

  23. 23

    Cornell, W. D. et al. A second generation force field for the simulation of proteins, nucleic acids, and organic molecules. J. Am. Chem. Soc. 117, 5179–5197 (1995).

    CAS  Article  Google Scholar 

  24. 24

    Santosh, M. & Maiti, P. K. Structural rigidity of paranemic crossover and juxtapose DNA nanostructures. Biophys. J. 101, 1393–1402 (2011).

    CAS  Article  Google Scholar 

  25. 25

    Joshi, H., Dwaraknath, A. & Maiti, P. K. Structure, stability and elasticity of DNA nanotubes. Phys. Chem. Chem. Phys. 17, 1424–1434 (2014).

    Article  Google Scholar 

  26. 26

    Maiti, P. K., Pascal, T. A., Vaidehi, N., Heo, J. & Goddard, W. A. Atomic-level simulations of seeman DNA nanostructures: the paranemic crossover in salt solution. Biophys. J. 90, 1463–1479 (2006).

    CAS  Article  Google Scholar 

  27. 27

    Sabharanjak, S. & Mayor, S. Folate receptor endocytosis and trafficking. Adv. Drug Deliv. Rev. 56, 1099–1109 (2004).

    CAS  Article  Google Scholar 

  28. 28

    Furey, W. S. et al. Use of fluorescence resonance energy transfer to investigate the conformation of DNA substrates bound to the Klenow fragment. Biochemistry 37, 2979–2990 (1998).

    CAS  Article  Google Scholar 

  29. 29

    Erben, C. M., Goodman, R. P. & Turberfield, A. J. Single-molecule protein encapsulation in a rigid DNA cage. Angew. Chem. Int. Ed. 45, 7414–7417 (2006).

    CAS  Article  Google Scholar 

  30. 30

    Bhatia, D., Chakraborty, S., Mehtab, S. & Krishnan, Y. A method to encapsulate molecular cargo within DNA icosahedra. Methods Mol. Biol. 991, 65–80 (2013).

    CAS  Article  Google Scholar 

  31. 31

    Lakshminarayan, R. et al. Galectin-3 drives glycosphingolipid-dependent biogenesis of clathrin-independent carriers. Nat. Cell Biol. 16, 595–606 (2014).

    CAS  Article  Google Scholar 

  32. 32

    Delacour, D. et al. Apical sorting by galectin-3-dependent glycoprotein clustering. Traffic 8, 379–388 (2007).

    CAS  Article  Google Scholar 

  33. 33

    Römer, W. et al. Shiga toxin induces tubular membrane invaginations for its uptake into cells. Nature 450, 670–675 (2007).

    Article  Google Scholar 

  34. 34

    Römer, W. et al. Actin dynamics drive membrane reorganization and scission in clathrin-independent endocytosis. Cell 140, 540–553 (2010).

    Article  Google Scholar 

  35. 35

    Renard, H.-F. et al. Endophilin-A2 functions in membrane scission in clathrin-independent endocytosis. Nature 517, 493–496 (2015).

    CAS  Article  Google Scholar 

  36. 36

    Johannes, L. & Popoff, V. Tracing the retrograde route in protein trafficking. Cell 135, 1175–1187 (2008).

    CAS  Article  Google Scholar 

  37. 37

    Kusumi, A., Ike, H., Nakada, C., Murase, K. & Fujiwara, T. Single-molecule tracking of membrane molecules: plasma membrane compartmentalization and dynamic assembly of raft-philic signaling molecules. Semin. Immunol. 17, 3–21 (2005).

    CAS  Article  Google Scholar 

  38. 38

    Kusumi, A., Sako, Y. & Yamamoto, M. Confined lateral diffusion of membrane receptors as studied by single particle tracking (nanovid microscopy). Effects of calcium-induced differentiation in cultured epithelial cells. Biophys. J. 65, 2021–2040 (1993).

    CAS  Article  Google Scholar 

  39. 39

    Wang, B., Kuo, J. & Granick, S. Bursts of active transport in living cells. Phys. Rev. Lett. 111, 1–16 (2013).

    Google Scholar 

  40. 40

    Bálint, Š., Verdeny Vilanova, I., Sandoval Álvarez, Á. & Lakadamyali, M. Correlative live-cell and superresolution microscopy reveals cargo transport dynamics at microtubule intersections. Proc. Natl Acad. Sci. USA 110, 3375–3380 (2013).

    Article  Google Scholar 

  41. 41

    Zajac, A. L., Goldman, Y. E., Holzbaur, E. L. F. & Ostap, E. M. Local cytoskeletal and organelle interactions impact molecular-motor-driven early endosomal trafficking. Curr. Biol. 23, 1173–1180 (2013).

    CAS  Article  Google Scholar 

  42. 42

    Lakadamyali, M., Rust, M. J. & Zhuang, X. Ligands for clathrin-mediated endocytosis are differentially sorted into distinct populations of early endosomes. Cell 124, 997–1009 (2006).

    CAS  Article  Google Scholar 

  43. 43

    Mayor, S. & Maxfield, F. R. Insolubility and redistribution of GPI-anchored proteins at the cell surface after detergent treatment. Mol. Biol. Cell. 6, 929–944 (1995).

    CAS  Article  Google Scholar 

Download references


The authors thank S. Surana and T. Pons for discussions and suggestions. The imaging facilities CIFF at NCBS and PICT-IBiSA / Nikon Imaging Centre at the Institut Curie-CNRS and the France-BioImaging infrastructure (ANR-10-INSB-04) are acknowledged. D.B. thanks CEFIPRA for the Charpak Fellowship, the Institute Curie and HFSP for postdoctoral fellowships. This work was supported by the following grants: 4803-B from CEFIPRA to B.D. and Y.K.; RGP0029/2014 from HFSP to Y.K. and L.J.; Agence Nationale pour la Recherche ANR-09-BLAN-283 from FPGG to B.D. and L.J.; ANR-11 BSV2 014 03 to L.J.; project 340485 from the European Research Council to L.J.; UL1 TR000430 from the National Center for Advancing Translational Sciences of the NIH and start-up support from the University of Chicago to Y.K. The Johannes team is members of Labex CelTisPhyBio (11-LBX-0038) and of Idex Paris Sciences et Lettres (ANR-10-IDEX-0001-02 PSL).

Author information




D.B. and Y.K. conceived the project and designed experiments; D.B. performed all experiments, some collaboratively. C.W., M.N., S.A., C.G. and B.N. contributed reagents and tools. S.A., V.P. and V.C. performed key experiments. H.J. and P.K.M. designed and performed molecular dynamics. D.B., C.W., P.K.M., L.J., B.D. and Y.K. designed experiments and analysed data. D.B., L.J. and Y.K. co-wrote the manuscript. All of the authors discussed the results and the manuscript.

Corresponding authors

Correspondence to Ludger Johannes or Benoit Dubertret or Yamuna Krishnan.

Ethics declarations

Competing interests

B.D. is also associated with NexDot, a for-profit company that provided the QDs used in this collaborative study.

Supplementary information

Supplementary information

Supplementary information (PDF 4817 kb)

Supplementary Movie 1

Supplementary Movie 1 (AVI 27483 kb)

Supplementary Movie 2

Supplementary Movie 2 (AVI 15032 kb)

Supplementary Movie 3

Supplementary Movie 3 (AVI 9705 kb)

Supplementary Movie 4

Supplementary Movie 4 (AVI 49367 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Bhatia, D., Arumugam, S., Nasilowski, M. et al. Quantum dot-loaded monofunctionalized DNA icosahedra for single-particle tracking of endocytic pathways. Nature Nanotech 11, 1112–1119 (2016). https://doi.org/10.1038/nnano.2016.150

Download citation

Further reading


Find nanotechnology articles, nanomaterial data and patents all in one place. Visit Nano by Nature Research