Masson's Trichrome Stain

Intended use:

Masson's Ttrichrome Stain selectively stains collagen fibers. Differentiation staining of collagen fibers and muscle fibers is an important diagnostic tool in pathological diagnosis.

Principle:

This method is modified from Mallory’s Trichome stain, which uses the mixture of two or three anion dyes or its function in sequence to stain the specimen. Staining results can be affected by the molecular weight of anion dyes and tissue permeability.

Demonstration of different tissue components can be made from selection of anion dyes with different molecular weight and tissue permeability.

Methods:

1. Fix tissues in Bouin’s Solution or Zenker’s Solution, wash in running tap water overnight, then dehydrate and embed.

2. Deparaffinize slides before washing with deionized water.

3. Stain in Weigert’s Iron Hematoxylin Solution for 5~10 minutes. Wash briefly in running tap water.

4. Place tissue in 1% hydrochloric acid ethanol. Wash in running tap water for a few minutes.

5. Stain in Ponceau-Acid Fucshin for 5~10 minutes. Rinse briefly in deionized water.

6. Place slides in 1% Phosphomolybdic Acid Solution for 5 minutes, then in Aniline Blue Solution for 5 minutes.

7. Place slides in 1% Acetic Acid for 1 minute, followed by dehydration in  95% alcohol for several times.

8. Dehydrate through 100% alcohol, clear in xylene and cover with mounting media.

Specifications:

Precaution:

1. Weigert’s Iron Hematoxylin solution should be prepared by mixing an equal part of Solution A and Solution B prior to use. Do not mix beforehand or else it will be oxidized and precipitated, resulting in the loss of staining capability.

2. to form precipitates and result in bad staining effect.

3. Tissues should be fixed in Bouin’s Solution or Zenker’s Solution. If fixed in 10% formaldehyde, first deparaffinize slides to deionized water, place in Bouin' s Solution overnight or incubate at 37℃for 1~2 hours, and then wash in running tap water (until yellow color disappears) before staining.

4. The length of treatment with 1% Phosphomolybdic Acid Solution can be determined under the microscope. The treatment can be stopped when muscle fibers appear red and collagen fibers pink.

Expected Results: