Grocott's Methenamine Silver

Classification: microorganism stain 

Purpose: stain fungi 

Mechanism of staining: induced argentaffin metallic impregnation 

Positive controls: any tissue containing fungi

Grocott's Methenamine Silver

Protozoa in lung




REAGENT

REAGENT

PURPOSE

MECHANISM OF STAINING

SOURCE OF ERROR

Chromic acid

Oxidization

Strong oxidizer that over-oxidizing all tissues, creating reactive aldehyde groups in fungi

Omitted: No aldehyde groups will be produced and no fungi demonstrated.

Too short: Background staining.

Too long: Over oxidation of aldehyde groups and no demonstration of fungi.

Sodium bisulphite

Bleaching

Removes brown discoloration created by the chromic acid

Omitted: Discoloration in counterstain

Too short: No effect.

Too long: No effect.

Methenamine silver (preheated to 60 Celsius)

Impregnation

Silver binds with the aldehyde groups. Aldehydes reduce silver to visible black color.

Omitted: Fungi will not be demonstrated.

Too short: Patchy staining of fungi may be seen.

Too long: Dark fungi that obscure details necessary to differentiate species (septae)

Gold chloride

Toning

 

Substitute silver with gold by ion exchange

Removes any brown color in other tissue components

Omitted: Collagen will remain golden brown.

Too long: Production of red tones due to overtoning. Will start to remove silver staining.

Too short: No effects on the staining.

 

Sodium thiosulphate

Fixation

 Removes unreacted silver

Omitted: Causes non-specific blackening of tissue over time.

Too long: No effects on the staining.

Too short: Causes blackening of tissue over time.

Light green or Neutral red

Counterstain

 

Omitted: No other tissue components demonstrated.

Too long: Fungi obscured.

Too short: Other tissues stain lightly and no detail seen.

 

Well Stained Slide

Fungi: black outline with grey interior


Lung: gree
n



Fungi: black-brown 



Possilby over-toned


Special Considerations

No metal forceps should be used when working with this stain.

References

Officer B. HIML251 Lecture notes: Silver Stains: Grocott’s Methenamine Silver, Jone’s Methenamine Silver, and Masson Fontana, March 18, 2009