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Published September 2020 | Supplemental Material + Accepted Version
Journal Article Open

Acoustic biosensors for ultrasound imaging of enzyme activity


Visualizing biomolecular and cellular processes inside intact living organisms is a major goal of chemical biology. However, existing molecular biosensors, based primarily on fluorescent emission, have limited utility in this context due to the scattering of light by tissue. In contrast, ultrasound can easily image deep tissue with high spatiotemporal resolution, but lacks the biosensors needed to connect its contrast to the activity of specific biomolecules such as enzymes. To overcome this limitation, we introduce the first genetically encodable acoustic biosensors—molecules that 'light up' in ultrasound imaging in response to protease activity. These biosensors are based on a unique class of air-filled protein nanostructures called gas vesicles, which we engineered to produce nonlinear ultrasound signals in response to the activity of three different protease enzymes. We demonstrate the ability of these biosensors to be imaged in vitro, inside engineered probiotic bacteria, and in vivo in the mouse gastrointestinal tract.

Additional Information

© 2020 Springer Nature Limited. Received 18 August 2019; Accepted 12 June 2020; Published 13 July 2020. The authors thank Z. Sun, A. Shur and R. Murray for sharing the protocols and reagents used for the cell-free TX–TL system. TEM was done at the Beckman Institute Resource Center for Transmission Electron Microscopy at Caltech. This research was supported by the National Institutes of Health (NIH) (no. R01-EB018975) and Defense Advanced Research Projects Agency (no. W911NF-14-1-0111). A.L. was supported by a National Science Foundation (NSF) Graduate Research Fellowship (no. 1144469) and the Biotechnology Leadership Pre-doctoral Training Program in Micro/Nanomedicine (Rosen Bioengineering Center and NIH Training Grant no. 5T32GM112592-03/04). D.P.S. was supported by an NSF Graduate Research Fellowship (no. 1745301). D.M. was supported by the Human Frontier Science Program (no. LT000637/2016). Related research in the Shapiro Laboratory is supported by the Heritage Medical Research Institute, Burroughs Wellcome Career Award at the Scientific Interface, Pew Scholarship in the Biomedical Sciences and Packard Fellowship for Science and Engineering. Author Contributions: A.L. and M.G.S. conceived the study. A.L., Z.J. and S.P.N. designed and planned the experiments. A.L., Z.J., S.P.N., D.P.S., A.L-G., M.B.S. and D. Malounda conducted the experiments. Z.J., D.P.S. and D. Maresca wrote the MATLAB scripts for ultrasound imaging and data processing. A.L., Z.J. and M.G.S. analyzed the data. A.L., Z.J. and M.G.S. wrote the manuscript with input from all authors. All authors gave approval to the final version of the manuscript. The authors declare no competing interests.


Correction to: Nature Chemical Biology https://doi.org/10.1038/s41589-020-0591-0, published online 13 July 2020 In the version of this article originally published, there were errors in Fig. 4 and Extended Data Figs. 5 and 7. In Fig. 4f, labels "+ aTc" and "− atc" were on the wrong sides. "+ aTc" should be placed over the blue bar on the right-hand column, and "− aTc" should be on the left. In Extended Data Fig. 5b, the colormap accompanying the nonlinear ultrasound image was blank. Additionally, the label "ASGWT" on the left should be "ARGWT", and the orange and blue overbar colors should be swapped, with orange on the right and blue on the left. In Extended Data Fig. 7, the colormaps accompanying the nonlinear ultrasound images in b and c were blank, and the significance asterisks above the nonlinear data in d and e were shown as crossed squares. The errors have been corrected in the HTML and PDF versions of the paper. Published: 23 July 2020.

Attached Files

Accepted Version - nihms-1647019.pdf

Supplemental Material - 41589_2020_591_Fig10_ESM.webp

Supplemental Material - 41589_2020_591_Fig11_ESM.webp

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Supplemental Material - 41589_2020_591_MOESM1_ESM.pdf

Supplemental Material - 41589_2020_591_MOESM2_ESM.pdf

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Supplemental Material - 41589_2020_591_MOESM7_ESM.pdf

Supplemental Material - 41589_2020_591_MOESM8_ESM.pdf


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August 22, 2023
October 23, 2023