High Iron Diamine－Alcian Blue Stain

 

Principle:

O-sulfate esters are stained brown-black with HID generated “in-situ” by the oxidation of a mixture of hydrochlorides of N, N-dimethyl meta- and para-phenylene diamines with ferric chloride. In subsequent steps, sialic acid and carboxyl groups are stained aquamarine blue with Alcian Blue at pH 2.5.

 

Specifications:

Contents	25Tests	Components
Alcian blue solution（pH2.5）	1vial×20ml	Alcian blue
HID-solution A	1vial×100ml	3-Amino-N，N-dimethylaniline dihydrochloride
HID-solution B	1vial×20ml	3-Amino-N，N-dimethylaniline dihydrochloride
Ferric Chloride solution	1vial×10ml	ferric chloride

 

Working Solution Preparation:

HID-solution A…………….40ml (20ml)

HID-solution B…………….10ml (5ml)

Ferric Chloride……………..1.5ml (0.75ml)

Prepare the solution prior to use.

 

Methods:

1. Deparaffinize tissue in deionized water.

2.Place in working solution at room temperature for 18~24 hours.

3. Rinse in running tap water.

4. Stain in Alcian blue solution（pH2.5）for 10~20 minutes.

5. Briefly wash with water. (If necessary, counterstain nuclei to fix the red color.)

6. Dehydrate, clear and mount with mounting media by following routine procedure.

 

Expected Results:

Acid Sulfmucus………………………..Purple to Dark brown

Carboxylated Acid Mucus…………….Blue

Nuclei…………………………………..Red (Counterstain nuclei to fix the red color.)

 

Precaution:

1. Prepare high iron diamine solution prior to use, and throw it off after use. The solution can be used once or twice if well stored in a refrigerator. However, the background and specificity may be influenced and changed.

2. Place smear in working solution at room temperature between 20~25℃. At lower room temperature extend staining time.

3. Diamine salt is toxic. Avoid contact with skin.