1. Wash the sections in PBS 0.5% Triton-X 100 at room temperature for 10 min X 3
2. Incubate sections in PBS 0.3% Triton-X 100 at room temperature for 30 min
3. Incubate sections with PBS 10% serum, and 0.3% Triton-X 100 at 4 ° C overnight (serum choice is determined by the host species of the secondary antibody)
4. Incubate the 1st antibody: dilute the 1st antibody in PBS 5% serum with 0.3% Triton-X 100, incubate the sections with the 1st antibody at 4 ° C overnight
5. Wash the sections in PBS 0.3% Triton-X 100 at room temperature for 10 min X 4
6. Incubate the secondary antibody: dilute the secondary antibody in PBS 5% serum with 0.3% Triton-X 100, incubate the sections with the secondary antibody at room temperature for 60 min (DAPI or other dye can be added at this time)
7. Wash the sections in PBS 0.3% Triton-X 100 at room temperature for 10 min X 4 and mount the slides
For the ß-Gal staining:
After incubation of the 1st antibody, use a biotinylated anti-rabbit antibody (1: 200), incubate at room temperature for 1 h to amplify the signal then wash the sections in PBS 0.3% Triton-X 100 at room temperature for 10 min X 4. Then use Avidin conjugate A488 (1: 500 to 1: 1000) for signal detection.