Sample Type Required: Deparaffinized and re-hydrated tissue sections on positively charged slides.
Fixative: 10% Neutral Buffered Formalin
| Step | Reagent | Time | Technical Notes |
| 1 | 12% Acetic Acid Solution | Brief Rinse |
|
| 2 | Working Colloidal Iron Solution (10mL Stock Colloidal Iron, 18mL Distilled Water, and 12mL Glacial Acetic Acid) | 1 Hour | Stock Colloidal Iron can be purchased commercially.
|
| 3 | 12% Acetic Acid Solution | 3 Changes x 3 Minutes Each |
|
| 4 | Ferrocyanide-Hydrochloric Acid Solution (25mL Potassium Ferrocyanide, 2% and 25mL Hydrochloric Acid, 2%) | 20 Minutes |
|
| 5 | Running Tap Water | 5 Minutes |
|
| 6 | Nuclear Fast Red Solution | 5 Minutes |
|
| 7 | Running Tap Water | 1 Minute | If slides are not washed well after nuclear-fast red staining, clouding will result when they are placed in the alcohol. |
Expected Results:
- Acid Mucins and Sialomucins = Deep Blue
- Nuclei = Pink to Red
- Cytoplasm = Pink
Post Staining Procedure:
- Tissue sections must be counterstained to recognize the presence of positive staining fully. This can be achieved using nuclear fast red, which provides a pink background.
- After counterstain, tissue sections should be rinsed well in distilled water, dehydrated with 95% alcohol, followed by 100% alcohol, cleared with xylene (or xylene substitute), and cover-slipped.
