Planarian Immunohistochemistry

1. 2% HCl/Holt, 4˚C, 5 min

2. Carnoy’ solution, 4˚C, 2 hrs

3. MeOH, -20˚C, vigorous shake

4. 6% H₂O₂/MeOH, RT, O/N

5. 70% EtOH/Holt, 4˚C, 30 min

6. 50% EtOH/Holt, 4˚C, 30 min

7. 30% EtOH/Holt, 4˚C, 30 min

8. Triton-PBS, 4˚C, 30 min

9. Triton-PBS, wash

10. 10% goat serum/Triton-PBS, RT, 2 hr

11. Primary Ab/10% goat serum/Triton-PBS, 4˚C, O/N

- 1/40

- 1/100

- 1/500

- 1/1000

12. Triton-PBS, wash

13. Triton-PBS, RT, 1hr ´ 4

14. 10% goat serum/Triton-PBS, RT, 2 hr

15. Secondary Ab (AP or HRP conjugated) /10% goat serum/Triton-PBS, 4˚C, O/N

16. Triton-PBS, wash

17. Triton-PBS, RT, 1hr ´ 4

18. Chromogenic reaction (use HRP, not AP, and use small (not whole molecule) secondary antibody!)