Modified Steiner Using Chapman's Modification

EMS26119

Components

Zinc Formalin

Silver Nitrate, 1% Aqueous

Gum Mastic, 2.5%

Hydroquinone, 2%

Silver Nitrate, 0.04%, Aqueous

Fixation

10% Buffered Neutral Formalin

Section

4-6µm

Staining Procedure

1. Deparaffinise and re-hydrate tissues to distilled water.
2. Sensitize sections in Zinc Formalin at 60°C in a water bath for 15 minutes (cover jar loosely). Preheat this solution.
3. Rinse in distilled water, 3 times for 2 minutes each.
4. Place in Silver Nitrate, 1%, in a Coplin Jar. Place in microwave (800W); start and run for 80 seconds. Remove from microwave and allow to stand in hot silver solution for 20 minutes (cover jar loosely).
5. Rinse in distilled water 3 times for 2 minutes each.
6. Dehydrate in 2 changes of 95% ethanol followed by 2 changes of 100% ethanol.
7. Treat with Gum Mastic, 2.5%, for 5 minutes.
8. Allow to air dry for 1 minute.
9. Rinse in distilled water (2 changes). Slides may stand in distilled water while reducing solution is being made, or longer if necessary.
10. To prepare reducing solution, mix just before use:
    
    a. Gum Mastic, 2.5% 10.0ml
    b. Hydroquinone, 2% 25.0ml
    c. Ethanol, 100% 5.0ml
    
    Filter and then add:
    d. Silver Nitrate, 0.04% 2.5ml
    
    DO NOT filter this silver!
11. Reducing solution at 45°C in water bath for 5-30 minutes or until sections have developed satisfactorily with the spirochetes at desired intensity and the background light yellow. Avoid an intensely stained background by checking microscopically. DO NOT PREHEAT THIS SOLUTION.
12. Rinse in distilled water to stop reaction.
13. Dehydrate through graded alcohol's clear in, Xylene (not Xylene substitute) and mount with Permount.

Results

Note: Acid wash all glassware used with silver solutions!

References

Margeson, L.S. Chapman CM: Use of Zinc Formalin as a sensitizer in Silver Stains for Spirochetes. Journal of Histotechnology , 19:2 p. 135-138, 6-96.