Aniline Blue 

Protocol List

 

 

 

Aniline Blue C.I. 42780 (ws 42755)

 

Properties

UV-VIS range: 594-610

Appearance: Purple-Brown Powder

Dye Class: Aminotriarylmethane

 

Biological Applications: Widely used as a counter stain, collagen stain, connective tissue stain, and in bacteriological media. 

 

Commercial Applications: Limited use as an indicator, formerly used as a basic dye in textiles.

 

Note:  Aniline Blue is a mixture of water soluble Acid Blue 22 (CI 42755) and Acid Blue 93 (CI 42780) 

 

 

           

Gomoris 1-step Trichrome Stain

 

 

Preparation of Stain Solutions   

 

Trichrome Stain

    Chromotrope 2R 0.6gm, Aniline Blue 0.3gm,

    Glacial Acetic Acid 1.0 ml, Phosphotungstic acid 0.8gm,

    distilled water 100 ml.

 

Fixation : 10%NBF

 

Staining Procedure

  1. Deparafinize sections through xylene.
  2. Hydrate tissues through graded ethanol.
  3. Rinse sections in distilled water.
  4. Place in Bouin's solution (at 58 degrees C) for 10 minutes.
  5. Rinse in running water until the yellow fades from the sections.
  6. Place sections in Weigert's iron hematoxylin solution for 5 minutes. (see Acid Fuchsin Van Gieson Stain)
  7. Rinse sections in running tap water for 10 minutes.
  8. Place sections in Trichrome stain for 15 minutes.
  9. Place in 0.5% acetic acid water 2 minutes, 2 changes.
  10. Deydrate in graded alcohols..
  11. Clear through xylene and mount with a synthetic resin medium.Results  Muscle Fibers - red, Collagen - blue, Nuclei - blue/black, Erythrocytes - red.           

Mallory's Connective Tissue Stain  

 

 Preparation of Stain Solutions     

0.5 % Acid Fuchsin

    0.5 gm of Acid Fuchsin in 100 ml distilled water

Aniline Blue Solution

    0.5 gm Aniline Blue, 2 gm Orange G, 1gm phosphotungstic acid, dissolve in 100 ml distilled water.

 

Fixation: Zenker 

 Staining Procedure

1.      Deparafinize sections through xylene.

2.      Hydrate tissues through graded ethanol.

3.      Rinse sections in distilled water.

4.    Dezenkerize if sections are Zenker fixed (10 minutes iodine, 10 minutes hypo solution).

5.    Rinse in distilled water.

6.    Stain sections in Acid Fuchsin solution for 1-5minutes depending on intensity of stain desired.

7.    Wipe off excess dye from slides and immerse slides in the Aniline Blue solution for 20minutes to 1 hour.

8.    Transfer to 95% ethanol, several changes to remove excess dye.

9.    Dehydrate clear and mount with a synthetic resin. 

      

Results  

 

Collagenous fibers - blue

Nuclei - red

neuroglia fibers - red

myelin - yellow

ground substances of cartilage, bone, mucus, amyloid - blue

 

 

 

 

REFERENCES:

Lillie, R.D. 1977. H.J. Conn's Biological Stains, 9th edition.  Williams & Wilkins, Baltimore.  pp. 289-291.

Churukian, Charles J. 2000, Manual of the Special Stains Laboratory, 5th edition. University of Rochester, Rochester. pp. 79-80.

Gomori, G.: A rapid one-step trichrome stain.  American Journal of Clinical Pathology. 20: 661-664,1950.

Green, F.J. 1990. The Sigma-Aldrich Handbook of Stains, Dyes and Indicators. Aldrich Chemical Comnpany, Milwaukee. pp.96-97.

Clark, G. 1981. Staining Procedures, 4rth edition.  Williams & Wilkins, Baltimore. pp.117.

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