c-Fos and Arc Immunohistochemistry on Rat Cerebellum

Materials and Reagents

1. c-Fos (raised in mouse, 1:1,000 dilution) (Santa Cruz Biotechnology, catalog number: sc-8074 )
   
2. Arc (raised in rabbit, 1:1,000 dilution) (Synaptic Systems, catalog number: 156002 )
   
3. c-Fos IHC, Alexa antimouse Fluor 488 (Life Technologies, Invitrogen™, catalog number: 51663A )
   
4. Arc IHC, Cy3-conjugated donkey anti-rabbit secondary antibody with 1:500 dilution (Jackson Laboratory, catalog number: 88069 )
   
5. Euthasol (Virbac, catalog number: 710101 )
   
6. Vectashield (Vector laboratory, catalog number: H-1000 )
   
7. PFA powder
   
8. 0.1 M PBS (pH 7.4)
   
9. TritonX-100
   
10. NaHPO₄
    
11. NaH₂PO₄
    
12. Paraformaldehyde (PFA)
    
13. NaOH
    
14. NaCl
    
15. Sucrose
    
16. Eythlen glycol (RNAse free) (Sigma-Aldrich, catalog number: E9129 )
    
17. Glycerol (RNase free)
    
18. Phosphate buffer (PB) (0.2 M stock, pH 7.4) (Solution 1) (see Recipes)
    
19. 4% Paraformaldehyde (PFA) for transcardial perfusion (Solution 2) (see Recipes)
    
20. 0.9% Saline (Solution 3) (see Recipes)
    
21. 30% sucrose in 4% PFA used as cryoprotectant (Solution 4) (see Recipes)
    
22. Phosphate buffer saline (PBS) (Solution 5) (see Recipes)
    
23. Cryoprotectant (Solution 6) (see Recipes)
    
24. Normal donkey serum containing 0.25% Triton-100 (see Recipes)
    
25. Primary Antibody (see Recipes)
    
26. Secondary Antibody (see Recipes)
    

Equipment

1. Fume hood
   
2. Water tab
   
3. Rib cage
   
4. Scissors
   
5. Forceps
   
6. Clippers
   
7. Cannula
   
8. Vial
   
9. Parafilm
   
10. Refrigerator
    
11. Tinfoil
    
12. Coverslip
    

Procedure