Imaging of Neuronal Mitochondria Using a Serial Block-Face Scanning Electron Microscope

Take a trimmed, resin-embedded mouse brain tissue section containing the hippocampus.

The section is stained with heavy metals to enhance contrast under an electron microscope.

Mount the section onto an aluminum pin using cyanoacrylate glue.

Coat the block sides with colloidal silver paste to create a conductive path to the aluminum pin.

Examine the tissue block using a serial block-face scanning electron microscope (SBFSEM).

Within the SBFSEM, the ultramicrotome sequentially slices the block into ultrathin layers while a low-voltage electron beam scans the exposed surface.

A backscattered electron detector captures high-resolution 2D images, revealing mitochondria and surrounding cellular structures.

Using appropriate software, adjust the brightness and contrast to improve image clarity.

Align the sequential images to reconstruct a 3D volume of the brain tissue. This 3D reconstruction allows precise mapping of mitochondrial morphology, spatial distribution, and their relationships within neuronal compartments.

Trim the samples to the area of interest and mount them onto an aluminum pin using gelling cyanoacrylate superglue. Then coat the sample with colloidal silver paste around the s