FIGURE SUMMARY
Title

Propagating-path uniformly scanned light sheet excitation microscopy for isotropic volumetric imaging of large specimens

Authors
Ping, J., Zhao, F., Nie, J., Yu, T., Zhu, D., Liu, M., Fei, P.
Source
Full text @ J. Biomed. Opt.
Fig. 1

(a) Schematic of the propagational light sheet scan using VCM. The confocal range of the laser sheet rapidly moving along the x direction. (b) Control timing diagram of the VCM motion and rolling shutter of the camera. (c) Device photo of VCM mounted with objective. (d) Through the synchronization, a virtual confocal slit is formed to exclusively collect the fluorescence excited by the Gaussian sheet waist (Video 1), as shown in (e) and compared to conventional static light-sheet geometry in (f). (g), (h) The plane images acquired under PULSE mode and conventional mode, respectively. (Video 1, MPEG, 5.47 MB [URL: https://doi.org/10.1117/1.JBO.24.8.086501.1]).
Fig. 2

(a) The beam profile at different propagation position adjusted by ETL (top row) and VCM (bottom row). (b) The thickness of light sheet at different focusing position by ETL (blue line) and VCM (orange line).
Fig. 3

(a)–(d) The signals distribution of fluorescent beads acquired using 1.7-μm static sheet, 1.7-μm PULSE, 3-μm static sheet, and 8-μm static sheet, respectively. (e), (f) Comparison of peak and averaged intensities of six selected signals shown in (a)–(d). Scale bars: 50  μm in (a)–(d) and 3  μm in insets.
Fig. 4

(a), (b) The reconstructed xz planes of fluorescent beads imaged by a 1.7-μm thin light sheet under conventional and PULSE modes. (c), (d) Vignette high-resolution views of seven selected beads at different x positions in (a) and (b). (e) The variations of axial intensity profiles of the selected beads in (c) and (d). Scale bars: 200  μm in (a), (b) and 5  μm in (c), (d).
Fig. 5

(a) The 3-D reconstruction of cleared mouse spinal cord with neurons labeled by GFP (thy1-GFP). (b) Three reconstructed yz planes with 200  μm interval in the propagation direction (x), showing the uniform isotropic resolution spanning a wide view. These results are further compared with the images by 1.7 and 8  μm static sheets, as shown in (c), (d), respectively. Insets in (b)–(d), line intensity profiles of the neuron fibers, indicating the different resolving power of three methods. (e) PULSE imaging of a blood vessel-tagged zebrafish larval (Tg flk1: mCherry). (e1)–(e3), The xy, xz, and yz views of the anterior part of the zebrafish. (e4) The 3-D volume rendering of the zebrafish. Scale bars: 50  μm in (b)–(d) and 100  μm in (e).
Acknowledgments
This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ J. Biomed. Opt.
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