For Occludine staining :

Wash 1X with PBS

- Fixation : formalin 15 min at RT

- Wash 2X with PBS 1X

- Triton 0.5%, for 5min

- Wash 1X with PBS 1X, 5min

- Wash 3X with PBS/glycine 1X, 10min

- Blocking with GS 10% in IF (plus Fab#152 for 30 min)

- Primary AB : dilute 1/50 in GS 10% (plus Fab#152), at least 2 hrs or O/N (4 degree) (ask johanne for the antibody)

- Wash 3X with IF buffer, 20min.

- Secondary : use S705 1/50 (40 min)

- Wash 3X with IF buffer, 20min.

- Nuclear staining : propidium iodide

- Wash

- Antifade etc…

For golgi staining :

Wash 1X with PBS

- Fixation : formalin 15 min at RT

- Wash 1X with PBS 1X, 5min

- Wash 3X with PBS/glycine 1X, 10min

- Blocking with GS 10% in IF (plus Fab#152 for 30 min)

- Primary AB : GM130 diluted 1/100 in GS 10% (plus Fab#152), at least 2 hrs or O/N (4 degree) (ask johanne for the antibody)

- Wash 3X with IF buffer, 20min.

- Secondary : use goat antimouse FITC 1/100 (40 min) (ask johanne for the antibody)

- Wash 3X with IF buffer, 20min.

- Nuclear staining : propidium iodide

- Wash

- Antifade etc…