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13 March 2024 Targeted illumination in widefield microscopy achieves confocal-quality images of neurites
Yao Wang, Jia Fan, Samuel Chung
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Proceedings Volume PC12848, Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXXI; PC128480L (2024) https://doi.org/10.1117/12.3000544
Event: SPIE BiOS, 2024, San Francisco, California, United States
Abstract
In widefield fluorescence imaging, out-of-focus and scattered light emanating from the cell body frequently obscures nearby dim fibers and degrades their contrast. Scanning techniques can ameliorate this issue but are hindered by a slower imaging speed and higher cost. We dramatically reduce stray light in widefield imaging by directing illumination primarily to neuron fibers. We identify fibers through real-time iterative image processing and pattern illumination onto these fibers using a digital micromirror device in a standard widefield microscope. By illuminating bright cell bodies with minimal light and in-focus fibers with high light intensity, we diminish the background and enhance the fibers' visibility. This methodology retains a high imaging speed and remains cost-effective. Employing this targeted illumination strategy, we have achieved confocal quality imaging of complex neurons in anesthetized C. elegans, ex vivo mouse brain slices, and restrained zebrafish larvae.
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Yao Wang, Jia Fan, and Samuel Chung "Targeted illumination in widefield microscopy achieves confocal-quality images of neurites", Proc. SPIE PC12848, Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXXI, PC128480L (13 March 2024); https://doi.org/10.1117/12.3000544
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KEYWORDS
Light sources and illumination
Microscopy
Image quality
Image segmentation
Microscopes
Neurons
Structured optical fibers
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