Functional imaging of live Zebrafish using fluorescence lifetime optical projection tomography (Conference Presentation)

Natalie Andrews; Samuel Davis; Carys Hay; Sunil Kumar; Marie-Christine Ramel; Laurence Bugeon; James McGinty; Margaret J. Dallman; Paul M. W. French

doi:10.1117/12.2252721

24 April 2017 Functional imaging of live Zebrafish using fluorescence lifetime optical projection tomography (Conference Presentation)

Natalie Andrews, Samuel Davis, Carys Hay, Sunil Kumar, Marie-Christine Ramel, Laurence Bugeon, James McGinty, Margaret J. Dallman, Paul M. W. French

Author Affiliations +

Proceedings Volume 10068, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XV; 1006808 (2017) https://doi.org/10.1117/12.2252721
Event: SPIE BiOS, 2017, San Francisco, California, United States

Abstract

Current microscopy techniques are not optimal to image fluorescence in whole live animals. We present fluorescence lifetime optical projection tomography (FLIM OPT) applied to imaging enzyme activity in live transgenic zebrafish expressing Förster Resonance Energy Transfer (FRET) biosensors. OPT can be considered the optical equivalent to x-ray CT. Samples are rotated through 360 with images acquired at set intervals, and a back projection technique is applied to reconstruct the 3D image. It can be performed in transmission or fluorescence modes, allowing a wide range of visualisation techniques, including FLIM. Combination of OPT with FRET FLIM can therefore provide functional information in 3D. The optimal size range for OPT is mm-cm, which fills the size gap between confocal and MRI and is also the size range for zebrafish, making them an ideal model for imaging. Transgenic zebrafish expressing a Caspase 3 FRET biosensor were generated on the TraNac background (a transparent mutant) to provide live readouts of apoptosis. We have shown that using FLIM OPT we can detect changes in Caspase 3 activity in both embryo and adult Tg(Ubi:Caspase3biosensor) zebrafish. Apoptosis was induced using 25 Gy from a 137Cs source and post irradiation an increase in fluorescence lifetime was quantified in the head region indicative of biosensor cleavage and Caspase 3 activity. Though development of compressive sensing and multiplexed imaging with two imaging arms we have applied OPT and FLIM OPT to adult zebrafish, enabling us to quickly acquire datasets so the fish can be recovered and imaged longitudinally.

Conference Presentation

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Natalie Andrews, Samuel Davis, Carys Hay, Sunil Kumar, Marie-Christine Ramel, Laurence Bugeon, James McGinty, Margaret J. Dallman, and Paul M. W. French "Functional imaging of live Zebrafish using fluorescence lifetime optical projection tomography (Conference Presentation)", Proc. SPIE 10068, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XV, 1006808 (24 April 2017); https://doi.org/10.1117/12.2252721

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KEYWORDS

Luminescence

Fluorescence lifetime imaging

3D image reconstruction

Biosensors

Fluorescence resonance energy transfer

Optical tomography

Projection systems

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