GMs

Author: Kelsey Goldman, HTL (ASCP)

Last Update: 2 January 2024

Cite this Page! -> Goldman, Kelsey. “GMS.” HistologyOutlines.Com, 2January 2024. Accessed (today)

Special Stain

Purpose:

The Groccot Methenamine Silver (GMS) is an Oxidation Reduction style silver stain for the detection of Fungi. The GMS is considered one of the most sensitive stains for fungi. (Henwood)(Churukian).

Control Tissues:

Tissues containing any of the following: Candida, Pneumocystis carinii, Cryptococcus Histoplasmosis, Blastomycosis, Aspergillus, Mucormycosis (Grizzle)

Staining Pattern:

Fungi- Black
Mucins and Glycogen- Gray
Background- Pale Green (Suvarna et al.)

Biochemistry Theory:

The GMS is an example of the oxidation/ reduction class of silver staining ( sometimes called combination stains). The principal of the oxidation/ reduction silver stain is that a strong oxidizing agent ( in this case chromic acid) is introduced to the tissue. This creates reductive groups in the aldehydes of the fungal walls. When “clear” silver ions are applied to the tissue and interact with these reductive groups they are reduced and become metallic silver, which is dark and visible. The metallic silver falls in the area of the aldehydes on the fungal walls (Grizzle).

Troubleshooting:

Using the wrong or a weak oxidizer will lead to nonspecific staining of the reticulum and carbohydrates. Not oxidizing enough will not create enough aldehydes to get a clear silver reaction
Overstaining is silver will cause none specific silver deposits
Understanding in silver will cause some organisms not to be stained (Grizzle)

Additional References:

Churukian, Charles J. “Consistent Silver Methods for Demonstrating Basement Membranes, Reticulum, and Fungi.” Journal of Histotechnology, vol. 19, no. 3, 1996, pp. 211–17, https://doi.org/10.1179/his.1996.19.3.211.
Grizzle, William E. “Theory and Practice of Silver Staining in Histopathology.” Journal of Histotechnology, vol. 19, no. 3, 1996, pp. 183–95, https://doi.org/10.1179/his.1996.19.3.183.
Henwood, Anthony F. “Looks like Fungi but It Isn’t – Identifying Pseudo-Fungi.” Journal of Histotechnology, vol. 40, no. 2, Apr. 2017, pp. 40–45, https://doi.org/10.1080/01478885.2017.1325588.
Suvarna, S. Kim, et al., editors. Bancroft’s Theory and Practice of Histological Techniques. Eighth edition, Elsevier, 2019.

Bancroft's Grcott Methenamine Silver for Fungi

Reagents

4% Chromic Acid
1% Sodium Bisulfate
0.1% Gold Chloride
5% Sodium Thiosulfate
0.21% Silver Nitrate Stock A- Prepared Silver Nitrate diluted in water; will store for 3 months in refrigerator
Methenamine Silver Borate Stock B- 27g Methenamine, 3.8g Sodium Borate Dehydrate, in 1000 mL of deionized water
0.2% light green solution- make fresh

Protocol

1. Deparaffinize and rehydrate

2. Oxidize in 4% Chromic acid for 30 minutes

3.Wash in deionized water

4. Quickly dip into 1% Sodium Bisulfate

5.Wash well in deionized water

6.Place in 60C oven or water bath (THIS MUST NOR BECOME HOTTER THAN 60C) for 15-20 minutes. Check for "brown bag" staining. nothing should yet be black (unless it is overheated or overstained)

7.Rinse well in deionized water

8.Tone in 0.1% Gold Chloride for 5 seconds, then rinse again in deionized water

9.Dip in 5% Sodium Thiosulfate for 5 seconds

10.Rinse in running tap water

11.Counterstain in light green for 5-15 seconds

12.Dehyrate and clear