Ultrastructural Analysis of Demyelination in Mouse Spinal Cord Cross-Sections

Obtain chemically-fixed spinal cord tissue from mice induced with autoimmune demyelination at defined time points during disease progression.

The disease triggers autoreactive immune cells to attack myelin proteins, degrading the myelin sheath surrounding neuronal axons.

Oligodendrocyte precursor cells are recruited to the affected site, mature into oligodendrocytes, and reform the myelin sheath.

Rinse the tissue with a buffer to remove excess fixative, then treat it with osmium, which binds to myelin lipids and enhances contrast for imaging.

Apply heavy metal stains that bind to macromolecules and enhance contrast.

Dehydrate the tissue using increasing alcohol concentrations. Take the tissue embedded in resin and obtain ultrathin sections.

Using electron microscopy, measure the myelin sheath thickness to classify axons as demyelinating with a swollen, thicker sheath, demyelinated with no sheath, or remyelinating with a thin sheath.

Identify axonal damage characterized by reduced spacing between neurofilaments and swollen mitochondria or complete degeneration.

Analyze the progression of axonal damage and repair over time to assess disease progression.