Alcian blue is a special stain that is used to identify acid mucin and mucopolysaccharides.

Special stain is any staining method other than H&E that may be used to visualize tissue structures, elements that not identify by H&E.

Alcian blue can be used in histology specially for light microscopy in paraffin embedded tissue sections (Commonly samples obtain from GI tract) and resin sections.


In histology alcian blue staining method is used to identify acid mucin and mucopolysaccharides seen in mesotheliomas, blood vessel walls and early lesions of atherosclerosis.


Alcian blue is basic dye containing positively charged groups comprised with copper molecules that gives it the blue colour which form salt linkage with poly anions (sulfated and carboxylated acid mucin and acid mucopolysaccharides).

Different types of acid mucin can be identified by changing the ph of the stain and alcian blue stain normally prepared at ph- 2.5.

Alcian blue solution

Alcian blue stain is mainly comprised with 1% Alcian blue, Glacial acetic acid and distilled water.

For preparing of 100ml solution it should be used 0.5g of 1% Alcian blue dye powder, 3ml of glacial acetic acid and 97ml of distilled water. 

  • First of all, labelled the slide and bring it to the water, it means put in to descending order of alcohol jars ((95%, 80%,70%, 60%, alcohol solution) and finally to the distilled water jar.
  • Then stain with 1% Alcian blue solution for 15 – 30 minutes and after that rinse with distilled water.
  • Then treat with safranin or neutral red for few seconds.
  • Neutral red or safranine is used as counter stain.
  • Then quick rinse in 95% alcohol and wash with distilled water.
  • Section is dehydrated and clear with clearing agent (xylene).
  • Finally mount the slide with DPX without any air bubbles.

*Method of staining can vary from one laboratory to another.


  • Acid mucin        – Blue
  • Nuclei                 – Reddish pink
  • Collagen fibers  – Cherry red
  • Cytoplasm         – Pale yellow

Alcian blue stain is really important for detection of acid mucin and acid mucopolysaccharides in connective tissues & epithelial tissues.

Combine alcian blue – PAS technique is helped to distinguish between acid and neutral mucin which acid mucin stain in blue by alcian blue and neutral mucin stain magenta in colour by PAS.

Hyaluronic acid detection, diagnosis of salivary gland pleomorphic adenoma and mucoepidermoid adenoma also is done by using alcian blue staining.

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