Immunostaining of Interneurons in a Rat Hippocampal Section

Take a fixed rat hippocampal section containing an interneuron filled with biocytin, a biotin-conjugated molecule.

Next, add a solution containing blocking proteins and a non-ionic detergent to prevent non-specific binding and permeabilize neurons.

Incubate with a primary antibody targeting specific cannabinoid receptors on interneurons.

Wash with buffer to remove excess antibodies.

Then, add a streptavidin-conjugated fluorescent dye and a fluorophore-conjugated secondary antibody.

During incubation, streptavidin binds strongly to biocytin, allowing visualization of the biocytin-filled interneuron, while the secondary antibody targets receptor-bound primary antibodies on the interneurons.

Rinse with buffer, removing unbound molecules.

Now, mount the section using aqueous mounting media. Position a coverslip and seal it.

Using a fluorescence microscope, visualize the biocytin-filled interneuron and cannabinoid receptor expression, enabling analysis of the interneuron's morphology and receptor distribution.

Lock the tissues with 10% blocking serum for one hour at room temperature. Subsequently, incubate the sections in primary antibody CB1R at room temperature overnight, to identify