To visualize individual axons in the spinal cord of living mice investigators used a transgenic mouse line (GFP-S mice), in which a neuron-specific fragment of the Thy1 promoter drives cytoplasmic expression of high levels of green fluorescent protein (GFP) in subsets of neurons, including sensory neurons in the dorsal root ganglia (DRG).
Investigators also used a spontaneous dominant mutation in C57/BL6 mice known as 'Wallerian degeneration slow' (WLD-S), which substantially delays Wallerian degeneration. They were crossed with GFP-S mice to obtain heterozygous WLD-SxGFP-S mice and acute axonal degeneration (AAD) was largely absent in these crosses. An antibody which recognizes fragment scalpain-mediated spectrin proteolysis, showed strong immunoreactivity at both lesion borders around the peak of AAD but not before the onset of AAD.
Immunohistochemistry was performed on fixed spinal cords with the following antibodies: anti-spectrin degradation product (1:500, gift of T.C. Saido, RIKEN Brain Science Institute, Japan) and anti-neurofilament 150 (1:1,000, Chemicon).