Fourier Light-Field Microscopy: Design, Optimization, and Applications
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Liu, Wenhao
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Abstract
Visualizing diverse anatomical and functional traits that span many spatial scales with high spatiotemporal resolution provides insights into the fundamentals of biological systems. Light-field microscopy (LFM) has recently emerged as a scanning-free, scalable method allowing for highspeed, volumetric imaging ranging from single-cell specimens to the mammalian brain. However, LFM is far prevented from broader applications by the prohibitive reconstruction artifacts and severe computational cost due to its nonuniform axial sampling and laterally variant PSF. To address the challenge, in this thesis, we report Fourier LFM (FLFM), a system that processes the light-field information through the Fourier domain, substantially overcomes the drawbacks of LFM and realizes fast 3D imaging with enhanced spatial resolution and extended imaging depth. Starting from the basic principle of optics, we established a complete framework consisting of theory and algorithm for the light propagation, image formation, volume reconstruction, and system characterization of FLFM. We proposed the generic rules for system design and
optimization, following which we developed two FLFM systems optimized for fast, 3D, live
imaging on, respectively, subcellular activities of organelles and functional and morphological
change of whole organoids. Based on these works as a solid base, we first, in Aim1, introduce
simultaneous two-color Fourier light-field imaging. By equipping a filter array behind the MLA
and allocating elemental images of imaged samples into different channels, we redistribute
fluorescent emission among spatial, angular, and chromatic dimensions and enable simultaneously
sensing ultrafast biological signals from two channels, breaking the camera limit on dual-color
FLFM imaging with alternative illumination. Then, in Aim 2, we developed an algorithm to
accelerate the reconstruction speed of FLFM by reducing the iterations in 3D deconvolution with
projection estimation, compressing 80%~90% of the time cost of reconstructing a volume. In the
end, in Aim3, we employed this IDC-FLFM system equipped with an accelerated reconstruction
algorithm in diverse biomedical studies, such as interrogating the mechanism of cancer invasion
in mammary organoids, toxicity assay on drugs for cardiac treatment, gene screening on
cardiovascular development, and detecting neural connections in animal locomotion. The imaging
results demonstrated the imaging capacity of our system for high-throughput screening and fast
time-lapsed 3D recording of rapid morphological change and functional traits, promising FLFM a
potent tool for imaging diverse phenotypic and functional information spanning broad molecular,
cellular, and tissue systems.
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Date
2024-04-26
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Dissertation