Methyl Green Counterstain Protocol

 

 

 

Reagents

 

0.1M Sodium Acetate Buffer, pH4.2:

Sodium acetate, trihydrate (MW 136.1) ------ 1.36 g

Distilled water ----------------------------------- 100 ml

Mix to dissolve and adjust pH to 4.2 using concentrated glacial acetic acid

 

Methyl Green Solution (0.5%)

Methyl green (ethyl violet free from Sigma) ---- 0.5 g

0.1M Sodium acetate buffer, pH4.2 ------------- 100 ml

Mix to dissolve.

 

Staining Pattern

 

Nuclear

 

Suggested Use

 

Counterstaining for immunohistochemistry or other special stains.

 

Staining Procedure

 

1.  Sections to distilled water after IHC.

2.  Stain in methyl green solution for 5 minutes at room temperature (60 C may produce slightly stronger stain).

3.  Rinse in distilled water (sections will look blue).

4.  Dehydrate quickly through 95% alcohol (10 dips, sections turn green), 2 changes of 100% alcohol (10 dips each) (alcohol used for dehydration removes some of the stain).

5.  Clear in xylene or xylene substitute.

6.  Mount with resinous mounting medium.