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@PHDTHESIS{Jaques_THESIS_2020,
         author = {Jaques, Christian},
       projects = {Idiap},
          title = {Active Illumination and Computational Methods for Temporal and Spectral Super-Resolution Microscopy},
           year = {2020},
         school = {EPFL},
            doi = {10.5075/epfl-thesis-7844},
       abstract = {Light microscopy is a tool of paramount importance for biologists and has been constantly
improved for the past four centuries. Despite many recent developments, microscopy techniques
still require improvement, especially to reach better temporal and spectral resolutions.
In particular, many high-end microscopes favor mostly spatial resolution, at the expense of
the latter two types of resolution.

In this thesis, we present methods based on the use of active illumination and computational
algorithms to increase temporal and spectral resolutions of microscopes. Our methods aim
to provide users with the flexibility to chose, within a single instrument, which type of resolution
is to be favored based on the application at hand. More generally, our approach has
fundamental implications on the signal sensing procedure, allowing, for example, to mitigate
temporal aliasing in sequences of images.

Our first method performs temporal super-resolution imaging of monochrome scenes using
a hue-encoded shutter. By making use of an active multi-spectral illumination, temporal
information is encoded in the hue of the acquisitions. We characterize the method showing a
resolution improvement of 2.8 and an increase of frame-rate of a factor 3. We demonstrate the
applicability of our method to bright-field transmission microscopy by applying the method
to the beating heart of a zebrafish embryo. We then extend this method to fluorescence microscopy.
We add a temporal regularization term to make the method robust to fluorescent
labelings inhomogeneities. We present an application of the method to the beating heart of a
fluorescently-labeled zebrafish that emits fluorescent light of two different colors. Implementing
our method within a light-sheet microscope allows us to reconstruct 3D+time videos of
the beating heart at twice the acquisition frame-rate.

Our second method offers a way to perform temporal generalized sampling by computing
simultaneous inner products with the sampled signal. Similarly to the first method, we take
advantage of working with multiple illumination hues to compute as many simultaneous
inner products, which we retrieve via an unmixing procedure. We use equivalent basic and
dual B-splines representations to ensure having finite-length and positive pre-filters, as well
as finite-support reconstruction functions. We show applications of our method to a fast
rotating target, as well as to the beating heart of a zebrafish embryo, both in transmission and
fluorescence microscopy.

Finally, we introduce a method to perform spectral imaging of repeating processes, such as
the beating heart. The method sequentially acquires multiple movies with various filters,
performs temporal registration of all movies and reconstructs a spectral movie through solving
of a spectral unmixing problem, pixel by pixel, at each time point. We characterize the method
and show a median error of approximately 10\%, by comparing reconstructions on a static
sample from our method with measurements obtained with a spectrometer. We then perform
validation by comparing static reconstructions with dynamic ones of the same sample. We
demonstrate the potential of the method to microscopy by performing spectral imaging of the
beating heart of a zebrafish embryo.

Taken together, these methods offer a versatile toolbox to improve the temporal or spectral
resolution in both bright field and fluorescence microscopy, which we foresee could be directly
implemented in a number of specialized instruments.},
            pdf = {https://publications.idiap.ch/attachments/papers/2020/Jaques_THESIS_2020.pdf}
}