JoVE Encyclopedia of Experiments
Neuroscience
0 views • 3:14 min • May 29th, 2025
Start with brain sections from a neurodegenerative mouse model.
Incubate with a detergent-supplemented blocking solution to increase tissue permeability and block non-specific binding. Discard the solution.
Introduce primary antibodies targeting amyloid-beta plaques. Incubate with agitation for antibodies to interact with amyloid-beta plaques, then wash.
Add red fluorophore-tagged secondary antibodies and incubate in the dark to allow the antibodies to interact.
Wash to remove unbound antibodies, followed by an alcohol wash.
Add curcumin, which binds beta-sheet-rich amyloids with high affinity, then wash.
Next, treat with increasing alcohol concentrations, dehydrating the brain sections. Add xylene to remove any traces of reagents.
Mount a section on a slide and observe under a fluorescence microscope.
Use a 590-nanometer wavelength laser to excite the red fluorophores on the secondary antibodies. Capture the image.
Switch to a 480-nanometer wavelength laser to excite the curcumin, emitting green fluorescence.
Overlay the images; red and green fluorescence colocalization confirms amyloid-beta plaque in the brain tissue.
For labeling of cryostat sectioned specimens with anti-A beta antibo
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