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Alcian Blue Stain For Acidic Mucins – Method and Tips

16 May

The alcian blue stain is this author’s preferred stain for the demonstration of acidic mucins. The dye was originally used for the dyeing of cotton before being discovered as a by Steedman in 1950.

The alcian blue itself is a cationic copper phthalocyanine dye which stains mucopolysaccharides and glycosaminoglycans a bluish colour. Within skin, acidic mucins can be found in many differing conditions such as a mucinoma, lupus and alopecia mucinosa.

Below is the preferred alcian blue method of this author

METHOD

Solutions

1g alcian blue in 3% acetic acid (check pH = 2.5)

1% safranin

1. Take sections to water

2. Cover slide with FILTERED alcian blue solution and leave for 20 minutes

3. Rinse in tap water

4. Counterstain with FILTERED 1% safranin for 10-15 seconds

5. Rinse in tap water

6. Dehydrate quickly, clear and mount.

Tips

– this author prefers an alcian blue staining time of about 20 minutes but can be done within the range of 10-30 minutes if desired.

– by reducing the pH to 0.2 the stainer can select for only strongly sulphated mucins. A pH of 1.0 stains both weak and strongly sulphated mucins. If using a lower pH method be sure no to rinse in tap water between the steps for too long as this can affect the alcian blue staining.

– this author prefers safranin as a counterstain due to its crisper staining, but safranin leeches out quickly in the dehydrating alcohols therefore blot dry after counterstaining and quickly dehydrate through the alcohols.

– neutral red can also be used as a counterstain.

– the alcian blue staining solution expires after approximately 6 months.

Thanks for reading and I welcome any comments and other tips.

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2 responses to “Alcian Blue Stain For Acidic Mucins – Method and Tips

  1. Dr Laurence SIFRE

    September 29, 2011 at 9:46 am

    Hi
    I read your post on Alcian blue and I encountered a problem with this stainng. My colours are quite light. Do you know a way to intensify them? Incubate Alcian blue longer? Concentrate it in the solution? Increase or decrease the pH?

    Have you any tips for me?

    Many thanks for your informations.

    Best regards
    Laurence

     
    • skinpathonline

      October 6, 2011 at 7:50 am

      Hi

      Sorry for my delayed response. I am assuming your staining of the mucins are lighter than expected (ie. lighter blue than expected). First of all I wouldn’t adjust the pH of the solution as this will lead to abberant staining and might lead to other unwanted areas being stained with the alcian blue. You can try to extend the alcian blue staining time, lets say to 30 mins but be careful as this can lead to greater background staining. You can also increase the concentration of the alcian blue in the solution by make sure the pH is remains as stated in the procedure. I have found if I increase the concentration of the alcian blue too much a precipitate forms on the slide.

      Let me know how you get along.

      Thanks.

      Paul.

       

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