Letter | Published:

Biogenic gas nanostructures as ultrasonic molecular reporters

Nature Nanotechnology volume 9, pages 311316 (2014) | Download Citation

Abstract

Ultrasound is among the most widely used non-invasive imaging modalities in biomedicine1, but plays a surprisingly small role in molecular imaging due to a lack of suitable molecular reporters on the nanoscale. Here, we introduce a new class of reporters for ultrasound based on genetically encoded gas nanostructures from microorganisms, including bacteria and archaea. Gas vesicles are gas-filled protein-shelled compartments with typical widths of 45–250 nm and lengths of 100–600 nm that exclude water and are permeable to gas2,3. We show that gas vesicles produce stable ultrasound contrast that is readily detected in vitro and in vivo, that their genetically encoded physical properties enable multiple modes of imaging, and that contrast enhancement through aggregation permits their use as molecular biosensors.

  • Subscribe to Nature Nanotechnology for full access:

    $59

    Subscribe

Additional access options:

Already a subscriber?  Log in  now or  Register  for online access.

References

  1. 1.

    et al. Use of diagnostic imaging studies and associated radiation exposure for patients enrolled in large integrated health care systems, 1996–2010. J. Am. Med. Assoc. 307, 2400–2409 (2012).

  2. 2.

    Gas vesicles. Microbiol. Rev. 58, 94–144 (1994).

  3. 3.

    Distribution, formation and regulation of gas vesicles. Nature Rev. Microbiol. 10, 705–715 (2012).

  4. 4.

    , & Ultrasound cardiography—contrast studies in anatomy and function. Radiology 92, 939–948 (1969).

  5. 5.

    , & Ultrasound microbubble contrast agents: fundamentals and application to gene and drug delivery. Annu. Rev. Biomed. Eng. 9, 415–447 (2007).

  6. 6.

    & Molecular imaging with targeted contrast ultrasound. Curr. Opin. Biotechnol. 18, 11–16 (2007).

  7. 7.

    & Clinical uses of microbubbles in diagnosis and treatment. Med. Biol. Eng. Comput. 47, 813–826 (2009).

  8. 8.

    et al. Neural progenitor cells labeling with microbubble contrast agent for ultrasound imaging in vivo. Biomaterials 34, 4926–4935 (2013).

  9. 9.

    , & Noninvasive cell-tracking methods. Nature Rev. Clin. Oncol. 8, 677–688 (2011).

  10. 10.

    et al. Nanoparticles as image enhancing agents for ultrasonography. Phys. Med. Biol. 51, 2179–2189 (2006).

  11. 11.

    et al. A novel site-targeted ultrasonic contrast agent with broad biomedical application. Circulation 94, 3334–3340 (1996).

  12. 12.

    et al. Hard shell gas-filled contrast enhancement particles for colour Doppler ultrasound imaging of tumors. Medchemcomm 1, 266–270 (2010).

  13. 13.

    , , , & Acoustic droplet vaporization for therapeutic and diagnostic applications. Ultrasound Med. Biol. 26, 1177–1189 (2000).

  14. 14.

    & A molecular imaging primer: modalities, imaging agents, and applications. Physiol. Rev. 92, 897–965 (2012).

  15. 15.

    Gasvakuolen, ein Bestandteil der Zellen der wasserblutenbildenden Phycochromaceen. Flora (Jena) 80, 241–282 (1895).

  16. 16.

    Pressure relationships of gas vacuoles. Proc. R. Soc. B 178, 301–326 (1971).

  17. 17.

    Harmonic imaging with ultrasound contrast agents. Clin. Radiol. 51, 50–55 (1996).

  18. 18.

    , & Basic acoustic properties of microbubbles. Echocardiography 19, 229–240 (2002).

  19. 19.

    , & in Molecular Imaging I (eds Semmler, W. & Schwaiger, M.) 223–245 (Springer, 2008).

  20. 20.

    , , , & Magnetic relaxation switches capable of sensing molecular interactions. Nature Biotechnol. 20, 816–820 (2002).

  21. 21.

    & Direct evidence for the role of light-mediated gas vesicle collapse in the buoyancy regulation of Anabaena flos-aquae (cyanobacteria). Limnol. Oceanogr. 29, 879–886 (1984).

  22. 22.

    , & Flotation characteristics of cyanobacterium Anabaena flos-aquae for gas vesicle production. Biotechnol. Bioeng. 60, 636–641 (1998).

  23. 23.

    , & Fate of bacteriophage lambda in non-immune germ-free mice. Nature 246, 221–222 (1973).

  24. 24.

    , , , & Advances in ultrasound biomicroscopy. Ultrasound Med. Biol. 26, 1–27 (2000).

  25. 25.

    & Acoustic modeling of shell-encapsulated gas bubbles. Ultrasound Med. Biol. 24, 523–533 (1998).

  26. 26.

    , & Oscillations of polymeric microbubbles: effect of the encapsulating shell. J. Acoust. Soc. Am. 107, 2272–2280 (2000).

  27. 27.

    Counting bubbles acoustically: a review. Ultrasonics 15, 7–13 (1977).

  28. 28.

    et al. Vascular permeability in a human tumor xenograft: molecular size dependence and cutoff size. Cancer Res. 55, 3752–3756 (1995).

  29. 29.

    , , & Nanoparticle PEGylation for imaging and therapy. Nanomedicine 6, 715–728 (2011).

  30. 30.

    & Gas vesicle genes identified in Bacillus megaterium and functional expression in Escherichia coli. J. Bacteriol. 180, 2450–2458 (1998).

  31. 31.

    , & Counting of gas vacuoles by electron microscopy in lysates and purified fractions of Microcystis aeruginosa. Protoplasma 73, 329–335 (1971).

  32. 32.

    & Average thickness of the gas vesicle wall in Anabaena flos-aquae. J. Mol. Biol. 129, 279–285 (1979).

  33. 33.

    & Regulatory multidimensionality of gas vesicle biogenesis in Halobacterium salinarum NRC-1. Archaea 2011, 716456 (2011).

Download references

Acknowledgements

The authors thank P. Lum for ultrasound equipment and advice, R. Zalpuri and K. McDonald for assistance with electron microscopy, K-K. Park and P. Khuri-Yakub for assistance with hydrophone measurements, E. Chérin for input on in vivo experiments and the manuscript, and A. Bar-Zion for assistance with data analysis. M.G.S. acknowledges funding from the Miller Research Institute and the Burroughs Wellcome Career Award at the Scientific Interface. Other funding was provided by California Institute for Regenerative Medicine grant RT2-02022 (D.V.S.), National Institutes of Health grant R01EB013689 (S.M.C), the Canadian Institutes of Health Research (F.S.F.) and the Terry Fox Foundation (F.S.F.).

Author information

Author notes

    • Mikhail G. Shapiro

    Present address: Division of Chemistry and Chemical Engineering, California Institute of Technology, 1200 East California Boulevard, Pasadena, California 91125, USA

Affiliations

  1. Miller Research Institute, University of California at Berkeley, 2536 Channing Way, Berkeley, California 94720, USA

    • Mikhail G. Shapiro
  2. Department of Bioengineering, 306 Stanley Hall MC #1762, University of California at Berkeley, Berkeley, California 94720, USA

    • Mikhail G. Shapiro
    • , Patrick W. Goodwill
    • , David V. Schaffer
    •  & Steven M. Conolly
  3. Department of Molecular and Cell Biology, 142 LSA #3200, University of California at Berkeley, Berkeley, California 94720, USA

    • Mikhail G. Shapiro
  4. Department of Electrical Engineering and Computer Science, University of California at Berkeley, Berkeley, California 94720, USA

    • Arkosnato Neogy
    •  & Steven M. Conolly
  5. Sunnybrook Research Institute, University of Toronto, 2075 Bayview Avenue, Toronto, Ontario M4N 3M5, Canada

    • Melissa Yin
    •  & F. Stuart Foster
  6. Department of Medical Biophysics, University of Toronto, 610 University Avenue, Toronto, Ontario M4N 3M5, Canada

    • F. Stuart Foster
  7. Department of Chemical and Biomolecular Engineering, University of California at Berkeley, Berkeley, California 94720, USA

    • David V. Schaffer

Authors

  1. Search for Mikhail G. Shapiro in:

  2. Search for Patrick W. Goodwill in:

  3. Search for Arkosnato Neogy in:

  4. Search for Melissa Yin in:

  5. Search for F. Stuart Foster in:

  6. Search for David V. Schaffer in:

  7. Search for Steven M. Conolly in:

Contributions

M.G.S. conceived and directed the study, planned the experiments, prepared the specimens, collected, analysed and interpreted the data, and wrote the manuscript, with input from all other authors. P.W.G. designed and constructed the imaging instrument and accompanying signal processing software, and assisted with initial experiments. A.N. designed, constructed and optimized the imaging instrument and accompanying signal processing software. F.S.F. and M.Y. designed, performed and analysed the data from in vivo experiments. All authors provided input on the study and experimental design, data analysis, data interpretation and the manuscript.

Competing interests

F. Stuart Foster is a consultant to VisualSonics.

Corresponding author

Correspondence to Mikhail G. Shapiro.

Supplementary information

PDF files

  1. 1.

    Supplementary information

    Supplementary Information

Videos

  1. 1.

    Supplementary Movie 1

    Supplementary Movie 1

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nnano.2014.32

Rights and permissions

To obtain permission to re-use content from this article visit RightsLink.