Low-temperature resin embedding of the whole brain for various precise structures dissection

Area of Science:

• Neuroscience
• Biochemistry
• Microscopy

Background:

• Resin embedding and ultra-thin sectioning are standard for biological tissue imaging.
• Conventional methods degrade quenchable fluorescent signals and affect pH-insensitive dyes.

Purpose of the Study:

• To develop a novel embedding method preserving fluorescence and reducing background.
• To enhance imaging of precise structures and multi-color labeled tissues.

Main Methods:

• Developed HM20-T, a low-temperature chemical polymerization embedding method.
• Tested fluorescence preservation of green fluorescent protein (GFP) and tdTomato.
• Evaluated compatibility with various fluorescent dyes and retained immunoreactivity.

Main Results:

• HM20-T doubled the fluorescence preservation ratio for GFP-tagged presynaptic elements and tdTomato-labeled axons.
• The method is compatible with diverse fluorescent dyes like DyLight 488 conjugated Lycopersicon esculentum lectin.
• Brain tissues retained immunoreactivity after HM20-T embedding.

Conclusions:

• HM20-T effectively preserves multi-color fluorescence and immunoreactivity in biological tissues.
• This method facilitates detailed morphological characterization and circuit analysis of whole brains.