Alcian Blue/Periodic Acid Schiff

Question 1

What histochemical technique is used to determine the presence of CHO?

Answer 1

Alcian blue and Periodic Acid Schiff (PAS) techniques

These techniques allow for broad classification into acid mucin and neutral mucins plus glycogen.

Question 2

What is the function of Alcian blue in histochemical staining?

Answer 2

Alcian blue is a copper-containing dye that binds to acidic groups in CHO, producing a blue color

It interacts with negatively-charged acidic mucins, such as carboxylated and sulphated glycoproteins, through electrostatic attraction.

Question 3

What type of mucins does Alcian blue primarily stain?

Answer 3

Acid mucins

This includes carboxylated and sulphated glycoproteins.

Question 4

Describe the two chemical steps involved in the Periodic Acid Schiff (PAS) reaction.

Answer 4

1. Periodic acid oxidizes glycol groups in polysaccharides to generate aldehyde groups.
2. Aldehydes react with basic fuchsin in Schiff’s reagent to form a magenta color.

This reaction is essential for staining neutral mucins and glycogen.

Question 5

What is the principle behind the use of Alcian blue and PAS together?

Answer 5

Staining acidic mucins with Alcian blue first blocks them from reacting with PAS, allowing neutral mucins to be stained bright magenta.

This method highlights the distinction between acid and neutral mucins.

Question 6

What color is produced when both acid and neutral mucins are present?

Answer 6

Dark blue/purple staining color

This coloration results from the combination of Alcian blue and PAS staining.

Question 7

Can Alcian blue and PAS be applied independently?

Answer 7

Yes

Each stain can be used separately to identify different mucins.

Question 8

How does pH affect the staining of acid mucins with Alcian blue?

Answer 8

Varying the pH of the Alcian blue solution ionizes different acid mucins

At pH 0.2, only strongly sulphated mucins bind the dye; at pH 1, both weakly and strongly sulphated mucins react; at pH 2.5, all acid mucins stain.

Question 9

At what pH do only strongly sulphated mucins bind Alcian blue?

Answer 9

pH 0.2

This specific pH condition is crucial for selective staining.

Question 10

At what pH do all acid mucins stain with Alcian blue?

Answer 10

pH 2.5

This pH allows for the staining of all types of acid mucins.