Thionin Stain

Stock Solutions

  1. 1.0 M Sodium Acetate Solution:16.4g Sodium Acetate (anhydrous) dissolved in 200 ml distilled H2O.
  2. 1.0 M Glacial Acetic Acid: add12.2 ml Glacial Acetic Acid to 200 ml distilled H2O.
  3. 0.5% Thionin Solution: 0.5 gm thionin in 100 ml distilled H2O.

The above solutions are stock solutions. They have a fairly long shelf life. Thionin is hard to dissolve. Use a glass stirring rod or a stir bar. Do not use a metal rod to stir the thionin. Filter the thionin Solution.

Thionin Staining Solution:

  1. 180 ml distilled H2O
  2. 9 ml Sodium Acetate Solution
  3. 21 ml Acetic Acid Solution

This is the buffer Solution. Adjust pH to 4.3...then add

  1. 18 ml 0.5% thionin Solution.

Staining Protocol for Fixed Tissue

  1. Distilled H2O.....3-5 min
  2. Thionin Stain.....20 min
  3. Distilled H2O.....3-5 min
  4. Distilled H2O.....3-5 min
  5. 70% Ethanol.......5 min
  6. 70% Ethanol.......3 min
  7. 95% Ethanol.......3 min
  8. ABS Ethanol.......3 min
  9. Xylene............3 min
  10. Xylene............5 min

Coverslip slides wet with permount or microkit.

Staining Protocol for Fresh-Frozen Sections

  1. 70% Ethanol.....5 min
  2. 50% Ethanol.....1 min
  3. Distilled H2O….1 min
  4. Thionin Stain…10 min
  5. Distilled H20….30 sec
  6. Distilled H2O…30 sec
  7. 50% Ethanol.....30 sec
  8. 95% Ethanol.....30sec
  9. ABS Ethanol.....30 sec
  10. Xylene..............30 sec
  11. Xylene................3 min

Coverslip slides wet using permount or microkit.

 

Steve Milway

Last modified: Wednesday, December 17, 2003