IHC Counterstain - Methyl Green Solution

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Tony Rook
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IHC Counterstain - Methyl Green Solution

Please find the following media preparation method:

IHC Counterstain:

Methyl Green Solution

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

Counterstain 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.

Link Reference:
http://www.ihcworld.com/_protocols/counterstain_solutions/methyl_green.htm