Materials and Reagents

1. Syringe
2. Filter (Tisch Scientific, Millex, syringe-driven filter unit, 0.22 µm)
3. Paraffin slides 
4. Glass staining jar (IMEB, SKU:SD-09)
5. M.O.M Kit for detecting primary antibodies on mouse tissue (Vector Laboratories, Inc., catalog number: PK-2200)
   Note: This Kit includes blocking solution and diluent for dilution of both primary and secondary antibodies (stored at 4 °C).
6. Biotinylated goat anti-mouse IgG antibody (Vector Laboratories, catalog number: BA-9200, stored at 4 °C)
7. Vectastain Elite ABC HRP Kit (Vector Laboratories, catalog number: PK-6101, stored at 4 °C)
8. Biotinylated goat anti-rat IgG antibody (Vector Laboratories, catalog number: BA-9400, stored at 4 °C)
9. Rat anti-mouse CD8α antibody [AbD Serotec (now Bio-Rad Laboratories), catalog number: MCA2694, stored at -20 °C]
10. Normal goat serum [Sigma-Aldrich (now Millipore, Sigma), catalog number: G9023-10ML, stored at -20 °C]
11. Toluidine blue O (Sigma-Aldrich, catalog number: T3260-5G)
12. 0.1 M Phosphate buffer (AlboChemicals, SKU: B0787MV4W2)
13. Xylene
14. Ethanol
15. Methanol
16. H₂O₂
17. PBS
18. Permount (Fisher Scientific, catalog number: SP15-100)
19. Tris-HCl
20. Blocking solution (see Recipes)
21. Counterstain stock solution (see Recipes)
22. Counterstain working solution (see Recipes)
23. DAB solution (see Recipes)
    

Equipment

1. Beakers (Nalgene, catalog number: 1542E06)
2. Orbit shaker (Labline, catalog number: 3520)
3. Regular light microscope (Wetzlar, Leitz Orthoplan) 
4. Oven

Procedure