Methenamin Silver Method for Argentaffin Cells

Created by Kathleen Patrick, Modified on Mon, 11 Nov at 9:46 AM by Kathleen Patrick

EMS26481

 

Components:

Weigert's Iodine Solution

Sodium Thiosulfate 5% Aqueous

Silver Nitrate 5% Aqueous

Methenamine 3% Aqueous

Borate Buffer pH 7.8

Gold Chloride 0.1% Aqueous

Safranin O Solution 0.1%


Fixation:

Formalin, Bouin’s or other Formalin containing fixatives. Dichromates and mercury salts impair contrast between granules and background.


Sections:

Cut paraffin at 3-5µm sections.


Staining Procedures:

  1. Deparaffinise and hydrate slides to distilled water.
  2. Treat for 10 minutes in Weigert’s Iodine followed by bleaching for 10 minutes in Sodium Thiosulfate, 5%. Wash for 10 minutes in running water. Rinse in two changes in distilled water.
  3. Place section in chemically clean coplin jars containing the buffered methenamine-silver nitrate solution at room temperature and place in 60°C paraffin oven for 3 – 3.5 hours. (preheating the solution to 60°C reduces impregnation time by ½ to 1 hour). Rinse in distilled water.
    1. * prepare methenamine-silver nitrate by mixing:
      Silver Nitrate, 5% - 5ml
      Methenamine, 3% - 100ml
      For working buffered stain add 8 ml of Borate Buffer, pH 7.8 to 30ml of methenamine-silver nitrate. Use chemically clean glassware throughout.
  4. Tone for 10 minutes in Gold Chloride, 0.1%. Rinse in distilled water.
  5. Fix in Sodium Thiosulfate, 5% for 2 minutes. Wash in running water for 5 minutes
  6. Counterstain in Acetic Safranin O, 0.1% for 5 minutes.
  7. Dehydrate with Acetone, clear in Xylene and mount.


Stain Results:  

Argentaffin Cells

Black

Coarse connective tissue of the submucosa

Variable amount of blackening by 3-3.5 hour of impregnation

Granules of eosinophil leukocytes, nuclei, smooth muscle & surface epithelium

Show additional blackening after incubation beyond 3-3.5 hour.

Granules of mast cells

Remain red after nuclei & reticulum are blackened


References:

Clark, G, ed.: Staining Procedures, 3rd ed.: Baltimore: Williams & Wilkins Co., 1973, p. 161

Gomori, G.: Arch Path., 45:48, 1948

Burtner, H.J. and Lillie, R.D., Stain Tech., 24:225-7, 1949.

 

Was this article helpful?

That’s Great!

Thank you for your feedback

Sorry! We couldn't be helpful

Thank you for your feedback

Let us know how can we improve this article!

Select at least one of the reasons

Feedback sent

We appreciate your effort and will try to fix the article