Contrast and axial confinement enhancement in deep imaging via HiLo-based line-scanning temporal focusing microscopy

High-speed, optical-sectioning imaging is highly desired in biomedical studies, as most bio-structures and bio-dynamics are in three-dimensions. Compared to point-scanning techniques, line scanning temporal focusing microscopy (LSTFM) is a promising method that can achieve high temporal resolution while maintaining a deep penetration depth. However, the contrast and axial confinement would still be deteriorated in scattering tissue imaging. Here, we propose a HiLo-based LSTFM, utilizing structured illumination to inhibit the fluorescence background and, thus, enhance the image contrast and axial confinement in deep imaging. We demonstrate the superiority of our method by performing volumetric imaging of neurons and dynamical imaging of microglia in mouse brains in vivo.

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Scattering reduction by structured light illumination in line-scanning temporal focusing microscopy

Biomedical Optics Express ◽

10.1364/boe.9.005654 ◽

2018 ◽

Vol 9 (11) ◽

pp. 5654 ◽

Cited By ~ 5

Author(s):

Yi Xue ◽

Kalen P. Berry ◽

Josiah R. Boivin ◽

Dushan Wadduwage ◽

Elly Nedivi ◽

...

Keyword(s):

Structured Light ◽

Light Illumination ◽

Structured Light Illumination ◽

Temporal Focusing ◽

Line Scanning

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Overcoming tissue scattering in wide-field deep imaging by extended detection and computational reconstruction

10.1101/611038 ◽

2019 ◽

Author(s):

Yuanlong Zhang ◽

Tiankuang Zhou ◽

Xuemei Hu ◽

Hao Xie ◽

Lu Fang ◽

...

Keyword(s):

Multiphoton Microscopy ◽

Tissue Imaging ◽

Wide Field ◽

Temporal Focusing ◽

Tissue Scattering ◽

Deep Imaging ◽

Detection Mode ◽

Computational Reconstruction ◽

Imaging Depth

AbstractCompared to the golden technique of point‐scanning multiphoton microscopy, line‐scanning temporal focusing microscopy (LTFM) is competitive in high imaging speed while maintaining tight axial confinement. However, considering its wide‐field detection mode, LTFM suffers from shallow penetration depth as a result of crosstalk induced by tissue scattering. In contrast to the spatial filtering based on confocal slit detection, we propose the extended detection LTFM (ED‐LTFM), the first technique to extract signals from scattered photons and thus effectively extend the imaging depth. By recording a succession of line‐shape excited signals in 2D and reconstructing signals under Hessian regularization, we can push the depth limitation in scattering tissue imaging. We valid the concept with numerical simulations, and demonstrate the performance of enhanced imaging depth in in vivo imaging of mouse brains.

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