En bloc staining with hydroquinone treatment for block face imaging

Area of Science:

• Electron Microscopy
• Biotechnology
• Materials Science

Background:

• Three-dimensional (3D) ultrastructural reconstruction requires high contrast imaging in scanning electron microscopy (SEM).
• Traditional heavy metal staining enhances material contrast for backscattered electrons (BSEs) but can be time-consuming for large volumes.
• Hydroquinone (HQ) treatment has been shown to increase specimen conductivity and secondary electron (SE) generation.

Purpose of the Study:

• To investigate the effect of hydroquinone (HQ) treatment on contrast enhancement for block face imaging (BFI) using SEs.
• To determine if HQ treatment can reduce the total image acquisition time for large-volume 3D reconstruction.

Main Methods:

• Mouse liver tissues were fixed and processed using a reduced-OTO staining protocol.
• Specimens were treated with hydroquinone (HQ) solution or used as controls.
• Stained specimens were embedded in epoxy resin, sectioned, coated with carbon, and observed using SEM.

Main Results:

• HQ-treated specimens exhibited increased secondary electron (SE) generation compared to controls.
• HQ treatment did not significantly increase backscattered electron (BSE) numbers.
• The contrast-to-noise ratio of BFI using SEs was improved with HQ treatment.

Conclusions:

• Hydroquinone (HQ) treatment enhances SE generation efficiency, improving contrast for SEM-based 3D reconstruction.
• HQ treatment offers a potential method to reduce image acquisition time for large-volume ultrastructural analysis.
• This technique is valuable for advancing 3D reconstruction in biological and materials science research.