Genetically Encoded Phase Contrast Agents for Digital Holographic Microscopy
Quantitative phase imaging and digital holographic microscopy have shown great promise for visualizing the motion, structure, and physiology of microorganisms and mammalian cells in three dimensions. However, these imaging techniques currently lack molecular contrast agents analogous to the fluorescent dyes and proteins that have revolutionized fluorescence microscopy. Here we introduce the first genetically encodable phase contrast agents based on gas vesicles. The relatively low index of refraction of the air-filled core of gas vesicles results in optical phase advancement relative to aqueous media, making them a "positive" phase contrast agent easily distinguished from organelles, dyes, or microminerals. We demonstrate this capability by identifying and tracking the motion of gas vesicles and gas vesicle-expressing bacteria using digital holographic microscopy, and by imaging the uptake of engineered gas vesicles by mammalian cells. These results give phase imaging a biomolecular contrast agent, expanding the capabilities of this powerful technology for three-dimensional biological imaging.
© 2020 American Chemical Society. Received: July 31, 2020; Revised: October 21, 2020; Published: October 29, 2020. A.F. was supported by the NSERC graduate fellowship. Research in the Shapiro lab was supported by the National Institutes of Health (R01EB018975), the Packard Fellowship in Science and Engineering, the Pew Scholarship in the Biomedical Sciences, and the Heritage Medical Research Institute. The Nadeau lab was supported by the Gordon and Betty Moore Foundation (Grant 4038) and the National Science Foundation (NSF) 1828793. Portions of this work were supported under a contract from the Jet Propulsion Laboratory (JPL), California Institute of Technology, under a contract with the National Aeronautics and Space Administration (NASA). The authors thank Raymond Bourdeau for assistance with initial experiments, Kurt Liewer for the hologram simulation code, and Dina Malounda for providing some of the samples. Author Contributions: A.F. and M.B. contributed equally. The authors declare no competing financial interest.
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