First day

1. Rinse the brain slices in PBS 1X 3x10 minutes on the shaker (Skip this step if the brain slices were store in PBS and not cryoprotectant)
2. Incubate the slices at room temperature with 5% of Donkey Serum in PBS-T 0.5% for about an hour on the shaker.
3. Incubate the slices at 4°C overnight on a shaker (one in the Gereau lab deli fridge) with 1:1000 of chicken anti GFP antibody (Aves Lab antibody in the -20°C antibody freezer)

Note : The antibody works very VERY well at 1:1000, it should probably work at 1:2000 dilution)

Second day

1. Rinse the brain slices in PBS 1X 3x10 minutes on the shaker
2. Incubate the slices at room temperature with 1:500 Goat anti chicken secondary antibody for 3 hours (Covered in aluminum foil)
3. Rinse the brain slices in PBS 1X 3x10 minutes on the shaker
4. Mount the brain slices

Example :