JoVE Encyclopedia of Experiments
Neuroscience
0 views • 1:23 min • March 4th, 2025
Take a rehydrated section of diseased human brain tissue containing Lewy bodies, or LBs.
Incubate with hydrogen peroxide solution to inhibit endogenous peroxidases and prevent non-specific staining.
Heat the section in citrate buffer to unmask alpha-synuclein epitopes in the LBs.
Apply a blocking agent to prevent non-specific antibody binding.
Remove the excess blocking agent and incubate the section with primary antibodies targeting alpha-synuclein.
Wash off unbound antibodies.
Apply biotinylated secondary antibodies that bind to the primary antibodies.
Remove excess antibodies.
Add streptavidin-biotin-peroxidase complexes, which bind to the biotinylated secondary antibodies.
Wash off unbound complexes.
Apply a color-producing substrate that reacts with the peroxidase, forming brown precipitates.
Remove the excess substrate.
Treat with a counterstain to color the nuclei blue, enhancing the contrast against the brown-stained alpha-synuclein within the LBs in the neuronal cytoplasm under a microscope.