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Detail of Fig6_Whole_brain_imaging166_20180328_221446

Kaoru Seiriki, Atsushi Kasai, Takanobu Nakazawa, Misaki Niu, Yuichiro Naka, Masato Tanuma, Hisato Igarashi, Kosei Yamaura, Atsuko Hayata-Takano, Yukio Ago, Hitoshi Hashimoto (2019) Whole-brain block-face serial microscopy tomography at subcellular resolution using FAST., Nature protocols, Volume 14, Number 5, pp. 1509-1529

Published in 2019 May (Electronic publication in April 8, 2019, midnight )

• doi: 10.1038/s41596-019-0148-4
• pmid: 30962606
  

(Abstract) Here, we describe an optimized and detailed protocol for block-face serial microscopy tomography (FAST). FAST enables high-speed serial section fluorescence imaging of fixed brains at an axonal spatial resolution and subsequent image data processing. It renders brain-wide anatomical and functional analyses, including structural profiling of nuclear-stained brain at the single-cell level, cell-type-specific mapping with reporter animal brains and neuronal tracing with anterograde/retrograde labeling. Light-sheet fluorescence microscopy of cleared brains is advantageous in regard to imaging speed, but its spatial resolution is generally limited, whereas the opposite is true for conventional confocal microscopy. FAST offers a solution to overcome these technical limitations. This protocol describes detailed procedures for assembling the FAST hardware, sample preparation, imaging and image processing. A single imaging session takes as little as 2.4 h per mouse brain, and sample preparation requires 1 to several days, depending on pretreatments; however, multiple samples can be prepared simultaneously. We anticipate that FAST will contribute to unbiased and hypothesis-free approaches for a better understanding of brain systems.

• Animals
• Brain/diagnostic imaging[major]
• Brain Chemistry/physiology
• Imaging, Three-Dimensional/methods[major]
• Male
• Mice
• Mice, Inbred C57BL
• Microscopy, Fluorescence/methods[major]
• Molecular Imaging/methods[major]
• Tomography/methods[major]