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European Microscopy Congress 2020 Abstract Database

Open source optical gating technologies enable two-day phase-locked time-lapse 3D fluorescence imaging of the developing and beating zebrafish heart

Open source optical gating technologies enable two-day phase-locked time-lapse 3D fluorescence imaging of the developing and beating zebrafish heart

Session

DHA.3 - Machine assisted acquisition and analysis of microscopy data

Authors

Alex Drysdale (1), Dr Jonathan Taylor (1), Dr Chas Nelson (1)

Affiliations

1. University of Glasgow

Keywords

developmental biology, fluorescence microscopy, open hardware, optical gating, zebrafish heart

Abstract text

Here we present our latest advances in adaptive prospective optical gating technologies that allow ‘computationally frozen’ 3D stacks of the beating zebrafish heart to be acquired over two-day long experiments.
High-resolution, long-term fluorescence microscopy of living zebrafish hearts allows scientists to probe specific features of heart development, repair and, in the case of zebrafish, regeneration. However, the fast, constant beating of the heart and slow changes in shape, size and position during development create barriers to stable imaging. The use of drugs, electrical and optical pacing or temporarily stopping the heart have all been used to enable stable imaging; however, all of these approaches are directly effecting the biology of interest [1]. Here we present a suite of open access technologies (hardware and software) that solve these problems.
Adaptive prospective optical gating is a ‘smart microscopy’ technique that enables cardiac-phase-locked capture of fluorescence images [2,3,4]. Thus, this technology enables the collection ‘computationally frozen’ 3D stacks of the beating heart and time-lapse imaging over the course of days, even during times of dramatic developmental changes.
Here we present our latest implementations and algorithms for adaptive prospective optical gating. Our latest system is a fully open-source, plug-and-play implementation that can be integrated into a range of commercial microscopes bringing this technology to labs and facilities without expensive new set-ups or the technical expertise needed for custom microscopy systems.
This fully open source implementation includes a next-generation algorithm for adaptive prospective optical gating. Based on the Needleman-Wunsch alignment algorithm, famous for it’s importance in bioinformatics, our new cascading Needleman-Wunsch algorithm enables a more robust phase-lock and enables stable two-day long time-lapse microscopy of the developing zebrafish heart.
We also show how of ‘smart microscopy’ techniques such as adaptive prospective optical gating can be important in reducing phototoxicity and photodamage during in vivo imaging. Our technique has little to no effect on zebrafish physiology and a much reduced rate of photobleaching compared to similar fluorescence approaches.
Our experiments have already begun to show never before seen biology and led to the exploration of new models of development. We believe that open source optical gating technology we present here provides a significant tool for biologists studying the zebrafish cardiac system and will provide answers not possible with existing technologies.

References

[1] Taylor, Jonathan M. Frontiers in physiology 5 (2014), p. 481.
[2] Taylor, Jonathan M., et al. Journal of biomedical optics 16.11 (2011), p. 116021.
[3] Taylor, Jonathan M., John M. Girkin, and Gordon D. Love. Biomedical optics express 3.12 (2012), p. 3043-3053.
[4] Taylor, Jonathan M., et al. Nature communications 10.1 (2019), p. 1-15.