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Biomedical Nanotechnology pp 51–62Cite as

Real-Time Quantum Dot Tracking of Single Proteins

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 726))

Abstract

We describe a single quantum dot tracking method that can be used to monitor individual proteins in the membrane of living cells. Unlike conventional fluorescent dyes, quantum dots (fluorescent semiconductor nanocrystals) have high quantum yields, narrow emission wavelengths, and excellent photostability, making them ideal probes in single-molecule detection. This technique has been applied to study the dynamics of various membrane proteins including glycine receptors, nerve growth factors, kinesin motors, and γ-aminobutyric acid receptors. In this chapter, a basic introduction and experimental setup for single quantum dot labeling of a target protein is given. In addition, data acquisition and analysis of time-lapse single quantum dot imaging with sample protocols are provided.

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References

  1. Bruchez, M., Moronne, M., Gin, P., Weiss, S., and Alivisatos, A. P. (1998) Semiconductor nanocrystals as fluorescent biological labels. Science 281, 2013–2016.

    Article  CAS  Google Scholar 

  2. Chan, W. C. and Nie, S. (1998) Quantum dot bioconjugates for ultrasensitive nonisotopic detection. Science 281, 2016–2018.

    Article  CAS  Google Scholar 

  3. Alivisatos, P. (2004) The use of nanocrystals in biological detection. Nat. Biotechnol. 22, 47–52.

    Article  CAS  Google Scholar 

  4. Kim, S., Lim, Y. T., Soltesz, E. G., De Grand, A. M., Lee, J., Nakayama, A., et al. (2004) Near-infrared fluorescent type II quantum dots for sentinel lymph node mapping. Nat. Biotechnol. 22, 93–97.

    Article  CAS  Google Scholar 

  5. Rosenthal, S. J., Tomlinson, I., Adkins, E. M., Schroeter, S., Adams, S., Swafford, L., et al. (2002) Targeting cell surface receptors with ligand-conjugated nanocrystals. J. Am. Chem. Soc. 124, 4586–4594.

    Article  CAS  Google Scholar 

  6. Liu, W., Choi, H. S., Zimmer, J. P., Tanaka, E., Frangioni, J. V., and Bawendi, M. (2007) Compact cysteine-coated CdSe(ZnCdS) quantum dots for in vivo applications. J. Am. Chem. Soc. 129, 14530–14531.

    Article  CAS  Google Scholar 

  7. McBride, J., Treadway, J., Feldman, L. C., Pennycook, S. J., and Rosenthal, S. J. (2006) Structural basis for near unity quantum yield core/shell nanostructures. Nano Lett. 6, 1496–1501.

    Article  CAS  Google Scholar 

  8. Nirmal, M., Dabbousi, B. O., Bawendi, M. G., Macklin, J. J., Trautman, J. K., Harris, T. D., et al. (1996) Fluorescence intermittency in single cadmium selenide nanocrystals. Nature 383, 802–804.

    Article  CAS  Google Scholar 

  9. Scholl, B., Liu, H. Y., Long, B. R., McCarty, O. J., O’Hare, T., Druker, B. J., et al. (2009) Single particle quantum dot imaging achieves ultrasensitive detection capabilities for Western immunoblot analysis. ACS Nano 3, 1318–1328.

    Article  CAS  Google Scholar 

  10. Temirov, J. P., Bradbury, A. R., and Werner, J. H. (2008) Measuring an antibody affinity distribution molecule by molecule. Anal. Chem. 80, 8642–8648.

    Article  CAS  Google Scholar 

  11. Dahan, M., Levi, S., Luccardini, C., Rostaing, P., Riveau, B., and Triller, A. (2003) Diffusion dynamics of glycine receptors revealed by single-quantum dot tracking. Science 302, 442–445.

    Article  CAS  Google Scholar 

  12. Cui, B., Wu, C., Chen, L., Ramirez, A., Bearer, E. L., Li, W.-P., et al. (2007) One at a time, live tracking of NGF axonal transport using quantum dots. Proc. Natl Acad. Sci. USA. 104, 13666–13671.

    Article  CAS  Google Scholar 

  13. Rajan, S. S., Liu, H. Y., and Vu, T. Q. (2008) Ligand-bound quantum dot probes for studying the molecular scale dynamics of receptor endocytic trafficking in live cells. ACS Nano 2, 1153–1166.

    Article  CAS  Google Scholar 

  14. Courty, S., Luccardini, C., Bellaiche, Y., Cappello, G., and Dahan, M. (2006) Tracking individual kinesin motors in living cells using single quantum-dot imaging. Nano Lett. 6, 1491–1495.

    Article  CAS  Google Scholar 

  15. Bouzigues, C., Morel, M., Triller, A., and Dahan, M. (2007) Asymmetric redistribution of GABA receptors during GABA gradient sensing by nerve growth cones analyzed by single quantum dot imaging. Proc. Natl Acad. Sci. USA. 104, 11251–11256.

    Article  CAS  Google Scholar 

  16. Tada, H., Higuchi, H., Wanatabe, T. M., and Ohuchi, N. (2007) In vivo real-time tracking of single quantum dots conjugated with monoclonal anti-HER2 antibody in tumors of mice. Cancer Res. 67, 1138–1144.

    Article  CAS  Google Scholar 

  17. Lang, E., Baier, J., and Köhler, J. (2006) Epifluorescence, confocal and total internal reflection microscopy for single-molecule experiments: a quantitative comparison. J. Microsc. 222, 118–123.

    Article  CAS  Google Scholar 

  18. Yao, J., Larson, D. R., Vishwasrao, H. D., Zipfel, W. R., and Webb, W. W. (2005) Blinking and nonradiant dark fraction of water-soluble quantum dots in aqueous solution. Proc. Natl Acad. Sci. U.S.A. 102, 14284–14289.

    Article  CAS  Google Scholar 

  19. Zhang, L., Neves, L., Lundeen, J. S., and Walmsley, I. A. (2009) A characterization of the single-photon sensitivity of an electron multiplying charge-coupled device. J. Phys. B At. Mol. Opt. Phys. 42, 114011.

    Article  Google Scholar 

  20. Michalet, X., Fabien, P., Thilo, D. L., Maxime, D., Marcel, P. B., Alivisatos, A. P., et al. (2001) Properties of fluorescent semiconductor nanocrystals and their application to biological labeling. Single Mol. 2, 261–276.

    Article  CAS  Google Scholar 

  21. Crocker, J. and Grier, D. (1996) Methods of digital video microscopy for colloidal studies. J. Colloid Interface Sci. 179, 298–310.

    Article  CAS  Google Scholar 

  22. Rife, J. C., Long, J. P., Wilkinson, J., and Whitman, L. J. (2009) Particle tracking single protein-functionalized quantum dot diffusion and binding at silica surfaces. Langmuir 25, 3509–3518.

    Article  CAS  Google Scholar 

  23. Saxton, M. J. and Jacobson, K. (1997) Single-particle tracking: applications to membrane dynamics. Annu. Rev. Biophys. Biomol. Struct. 26, 373–399.

    Article  CAS  Google Scholar 

  24. Saxton, M. J. (2008) A biological interpretation of transient anomalous subdiffusion. II. Reaction kinetics. Biophys. J. 94, 760–771.

    Article  CAS  Google Scholar 

  25. Martin, D. S., Forstner, M. B., and Kas, J. A. (2002) Apparent subdiffusion inherent to single particle tracking. Biophys. J. 83, 2109–2117.

    Article  CAS  Google Scholar 

  26. Liu, W., Howarth, M., Greytak, A. B., Zheng, Y., Nocera, D. G., Ting, A. Y., et al. (2008) Compact biocompatible quantum dots functionalized for cellular imaging. J. Am. Chem. Soc. 130, 1274–1284.

    Article  CAS  Google Scholar 

  27. Warnement, M. R., Faley, S. L., Wikswo, J. P., and Rosenthal, S. J. (2006) Quantum dot probes for monitoring dynamic cellular response: reporters of T cell activation. IEEE Trans. Nanobioscience 5, 268–272.

    Article  Google Scholar 

  28. Warnement, M. R., Tomlinson, I. D., Chang, J. C., Schreuder, M. A., Luckabaugh, C. M., and Rosenthal, S. J. (2008) Controlling the reactivity of ampiphilic quantum dots in biological assays through hydrophobic assembly of custom PEG derivatives. Bioconjug. Chem. 19, 1404–1413.

    Article  CAS  Google Scholar 

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Acknowledgments

The authors wish to thank their colleagues in the group, especially Dr. James McBride, Dr. Michael Schreuder, Albert Dukes, and Oleg Kovtun, for all the helpful discussions and suggestions. We thank Dr. David Piston for helpful advice with single quantum dot tracking experimental setup. This work was supported by grants from National Institutes of Health (R01EB003778).

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Authors and Affiliations

  1. Department of Chemistry, Department of Pharmacology, Department of Chemical and Biomolecular Engineering, Department of Physics and Astronomy, Vanderbilt University, Nashville, TN, USA

    Sandra J. Rosenthal

Authors
  1. Jerry C. Chang
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  2. Sandra J. Rosenthal
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Corresponding author

Correspondence to Sandra J. Rosenthal .

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Editors and Affiliations

  1. Center for Nanoscale Materials, Argonne National Laboratory, S. Cass Avenue 9700, Argonne, 60439, Illinois, USA

    Sarah J. Hurst

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© 2011 Springer Science+Business Media, LLC

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Chang, J.C., Rosenthal, S.J. (2011). Real-Time Quantum Dot Tracking of Single Proteins. In: Hurst, S. (eds) Biomedical Nanotechnology. Methods in Molecular Biology, vol 726. Humana Press. https://doi.org/10.1007/978-1-61779-052-2_4

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  • DOI: https://doi.org/10.1007/978-1-61779-052-2_4

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-051-5

  • Online ISBN: 978-1-61779-052-2

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