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Southgate's Mucicarmine Staining Protocol
NovaUltra Special Stain Kits
Principle
Positively charged carmine-aluminium complex bonds with negatively charged acid mucins. The carmine solution is highly complex, and the detailed rationale is not fully understood.
Strongly sulphated mucins are variable in their reaction, neutral mucins staining not at all, whilst the other acidic mucins (particularly hyaluronic acid) stains strongly.
So as not to mask the staining result, counterstains which also demonstrate mucins (eg Ehrlich’s haematoxylin) are to be avoided.
Technical Points
(step 2) – do not use Ehrlich’s haematoxylin, as it stains mucins
Method
1. Bring sections to distilled water
2. Stain nuclei with alum haematoxylin 2 mins
3. Wash in running tap water
4. Differentiate in acid alcohol
5. Rinse in tap water
6. Blue in Scott’s tap water substitute
7. Wash in running tap water
8. Stain with mucicarmine solution 20 mins
9. Wash in running tap water
10. Dehydrate, clear and mount.
Results
mucins................................................. magenta
nuclei...................................................blue
cryptococcus neoformans.............................magenta
Reagent Formulae
1. alum haematoxylin (see Lillie Mayer)
2.
mucicarmine solution
carmine (CI 75470)
1.0 g
50% alcohol
100.0 ml
aluminium hydroxide
1.0 g
anhyd aluminium chloride 0.5 g
Grind the carmine to a fine powder, place in a 500 ml Ehrlenmeyer flask, add the 50% alcohol, and mix. Add the aluminium hydroxide, mix. Add the aluminium chloride, mix. Bring to the boil, and boil gently for two to three minutes. Cool and filter into a reagent bottle. Store at 4ºC. The solution keeps for six months or so.
References
Southgate HW,(1927),J.Path.Bact.,30,729