AB & ABPAS Flashcards
Which carbohydrates are in group 1?
Neutral polysaccharides
What carbohydrates are in group 2?
Acid mucopolysaccharides (anionic heteroglycans)
(all are acidic and attached to proteins)
What carbohydrates are in group 3?
Glycoproteins (mucins, mucoid, mucoprotein, mucosubstances)
What carbohydrates are in group 4?
Cerebrosides (lipid attached to carbs)
Phosphatides (non-carb containing lipids)
Which stains demonstrate mucin?
Mayer mucicarmine method (MMM or MUCIN)
Alcian Blue (AB)
Periodic Acid Schiff (PAS)
Colloidal iron
What is Alcian Blue used to demonstrate?
Acid mucins (Mucopolysaccharides) and Sialomucins (Glycoproteins)
What type of stain is Alcian Blue?
Carbohydrate stain
What are the target tissue components of Alcian Blue?
Group 2 and 3 Carbohydrates
What is the mechanism of staining of Alcian Blue?
Ionic bonding or salt linkage
What are the positive controls of Alcian Blue?
Small intestine, colon, appendix
Why is the duodenum the best control for ABPAS?
It has the Bruner’s gland that only picks up PAS
What is the most sensitive control for the viability of PAS staining reagents?
A small section of kidney
What are the characteristics of Alcian Blue?
It’s a copper phthalocyanin, a basic dye
It’s water soluble
Why is Alcian blue blue?
It’s blue because of copper
With what kind of solution is Alcian Blue made up of?
3% acetic acid solution
What is the consequence if you make the Alcian Blue solution to a pH of 1.0?
Only sulfated acid mucopolysaccharides and sulfated sialomucins (glycoproteins) will pick up the stain
What are the advantages of Alcian Blue over most cationic dyes?
Water, alcohol, weak acids, and other counterstaining solutions won’t extract the dye from stained sections.
The incubation time is adjustable.
It can be combined with PAS/Hx to demonstrate neutral and acid mucins.
What is the mechanism of staining of Alcian Blue?
Ionic bonding
What is the charge of the tissue and the dye ion when the tissue is acidophilic?
The tissue is positive
The dye ion is negative
What is the charge of the tissue and the dye ion when the tissue is basophilic?
The tissue is negative
The dye ion is positive
What is a key characteristic of an amphoteric dye?
It has groups that can change their charge based on the pH
How do acidic and basic dyes change as the pH changes?
As the pH increases, a basic dye will have a stronger reaction and thus impart a more vibrant colour. Conversely, an acidic dye will have a weaker reaction and impart less colour. The opposite is true when the pH decreases.
What type of dye have studies shown Alcian Blue more likely is?
An amphoteric dye
How does Alcian Blue behave at a pH of 1.0-4.0 and at a pH of 4.0-7.0, and what does this prove about this dye?
At a pH of 1.0-4.0 it will behave as a basic dye. At a pH of 4.0-7.0 it will behave as an acidic dye. This indicates it is an amphoteric dye.
What are the 8 steps of the Alcian Blue staining technique?
- 3% acetic acid
- Alcian blue solution
- 3% acetic acid
- RTW
- DW
- Nuclear fast red
- RTW
- DCM
What are the 12 steps of the ABPAS staining technique?
- 3% acetic acid
- Alcian blue solution
- RTW
- DW
- Periodic acid
- RTW
- DW
- Schiff’s reagent
- RTW
- Harris hematoxylin
- RTW
- DCM
What reagent is different when staining with AB at a pH of 1.0?
HCl is used instead of acetic acid
What is the purpose of the acetic acid rinse in the AB staining technique?
To protect the AB staining solution from pH changes due to the introduction of water.
At what pH is staining done with AB?
2.5 or 1.0
Is AB stained progressively or regressively?
Progressively and directly
Which components does AB attach to at pH 2.5 vs pH 1.0?
Carboxylated and sulphated negatively charged mucopolysaccharides and sialomucins at pH 2.5.
Sulphated negatively charged mucopolysaccharides and sialomucins only at pH 1.0.
Is the end product of AB staining soluble?
No
What type of stain the AB counterstain?
A nuclear stain
What staining principle does the AB counterstain use?
Ionic bonding
What is the most common counterstain used for AB?
Nuclear Fast Red
What is the AB staining result expectation? How do the results differ at a pH of 1.0 vs a pH of 2.5?
Mucin should be well stained and the counterstain should enhance the mucin staining.
At a pH of 1.0: sulphated mucopolysaccharides are pale blue. The background is pink to red
At a pH of 2.5: Weakly acidic mucopolysaccharides, hyaluronic acid and sialomucins are dark blue. The background is pink to red.
Why are goblet cells purple in ABPAS?
Because they pick up both stains
