“…S12 ). Compared to other LFM techniques which require scanning 33 or multiview imaging 22 , DiLFM gives an efficient performance-improving solution without any hardware modifications. Fig.…”
Light field microscopy (LFM) has been widely used for recording 3D biological dynamics at camera frame rate. However, LFM suffers from artifact contaminations due to the illness of the reconstruction problem via naïve Richardson–Lucy (RL) deconvolution. Moreover, the performance of LFM significantly dropped in low-light conditions due to the absence of sample priors. In this paper, we thoroughly analyze different kinds of artifacts and present a new LFM technique termed dictionary LFM (DiLFM) that substantially suppresses various kinds of reconstruction artifacts and improves the noise robustness with an over-complete dictionary. We demonstrate artifact-suppressed reconstructions in scattering samples such as Drosophila embryos and brains. Furthermore, we show our DiLFM can achieve robust blood cell counting in noisy conditions by imaging blood cell dynamic at 100 Hz and unveil more neurons in whole-brain calcium recording of zebrafish with low illumination power in vivo.
“…S12 ). Compared to other LFM techniques which require scanning 33 or multiview imaging 22 , DiLFM gives an efficient performance-improving solution without any hardware modifications. Fig.…”
Light field microscopy (LFM) has been widely used for recording 3D biological dynamics at camera frame rate. However, LFM suffers from artifact contaminations due to the illness of the reconstruction problem via naïve Richardson–Lucy (RL) deconvolution. Moreover, the performance of LFM significantly dropped in low-light conditions due to the absence of sample priors. In this paper, we thoroughly analyze different kinds of artifacts and present a new LFM technique termed dictionary LFM (DiLFM) that substantially suppresses various kinds of reconstruction artifacts and improves the noise robustness with an over-complete dictionary. We demonstrate artifact-suppressed reconstructions in scattering samples such as Drosophila embryos and brains. Furthermore, we show our DiLFM can achieve robust blood cell counting in noisy conditions by imaging blood cell dynamic at 100 Hz and unveil more neurons in whole-brain calcium recording of zebrafish with low illumination power in vivo.
“…Methods to improve signal extraction in scattering tissue have been demonstrated by computationally extracting fluorescence sources without reconstruction, 15,20,22,[38][39][40] although reconstruction-less signal extraction cannot resolve the propagation of calcium signals throughout spatially extended structures such as dendrites. Combining the principles of confocal microscopy with LFM, 41 selective-volume illumination, 19,42,43 These results demonstrate the capabilities and limitations of two light-field reconstruction algorithms for high SNR calcium fluorescence imaging. The trade-offs described above highlight the importance of adapting the volume reconstruction strategy to the scientific goals and requirements of future neurophysiology experiments.…”
Light field microscopy (LFM) enables fast, light efficient, volumetric imaging of neuronal activity with functional fluorescence indicators. Here we apply LFM to single-cell and bulk-labeled imaging of the red calcium dye, CaSiR-1 in acute mouse brain slices. We compare two common light field volume reconstruction algorithms: synthetic refocusing and Richardson-Lucy 3D deconvolution. We compare temporal signal-to-noise ratio (SNR) and spatial signal confinement between the two LFM algorithms and conventional widefield image series. Both algorithms can resolve calcium signals from neuronal processes in three dimensions. Increasing deconvolution iteration number improves spatial signal confinement but reduces SNR compared to synthetic refocusing.
“…Cardenas-Diaz et al were able to characterize candidate genes in vitro in the human EndoC-βH1 cell line by generating a dual reporter to express both an insulin-luciferase fusion protein and a GCaMP6s variant [112]. The slow kinetics, wide dynamic range, and high baseline brightness of GCaMP6s was suitable for studying Ca 2+ flux in β cells with fluorescence microscopy or pan-neuronal expression in mice and zebrafish [86,113]. [98].…”
Since the 1970s, the emergence and expansion of novel methods for calcium ion (Ca2+) detection have found diverse applications in vitro and in vivo across a series of model animal systems. Matched with advances in fluorescence imaging techniques, the improvements in the functional range and stability of various calcium indicators have significantly enhanced more accurate study of intracellular Ca2+ dynamics and its effects on cell signaling, growth, differentiation, and regulation. Nonetheless, the current limitations broadly presented by organic calcium dyes, genetically encoded calcium indicators, and calcium-responsive nanoparticles suggest a potential path toward more rapid optimization by taking advantage of a synthetic biology approach. This engineering-oriented discipline applies principles of modularity and standardization to redesign and interrogate endogenous biological systems. This review will elucidate how novel synthetic biology technologies constructed for eukaryotic systems can offer a promising toolkit for interfacing with calcium signaling and overcoming barriers in order to accelerate the process of Ca2+ detection optimization.
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