Immunohistochemical Staining of Nerve Fibers in the Nucleus Basalis of Meyner of the Mouse Brain

Take a fixed mouse brain section containing the nucleus basalis of Meynert, rich in nerve fibers expressing the enzyme choline acetyltransferase.

The section is pretreated to deactivate endogenous peroxidase.

Add a detergent to permeabilize cellular membranes, followed by a blocking solution to prevent non-specific antibody binding.

Replace the solution with primary antibodies that bind to choline acetyltransferase.

Wash with buffer to remove unbound antibodies. Incubate with biotin-tagged secondary antibodies, which bind to the primary antibodies.

Remove unbound antibodies and add avidin-biotin-peroxidase complexes, which bind to the secondary antibodies.

Wash to remove excess complexes.

Add a chromogenic substrate that reacts with peroxidase, forming a colored precipitate.

Rinse with distilled water to remove excess precipitate.

Transfer the section onto a gelatinized slide.

Next, dehydrate the section using increasing ethanol concentrations, followed by xylene.

Mount the section and observe under a microscope to visualize the stained nerve fibers.

For immunohistochemical labeling of the tissue sections, after blocking any nonspecific binding with 10% normal bovine serum in 0.1%