2-3. Modified Heidenhain's Azan (Ref 2)
Staining Procedures For Plastic Embedded Tissue
Verified at the Applications Laboratory of the Biomedical Division, Sorvall
Microtomes
SOLUTIONS:
| Solution A |
|
1.0g |
|
Azocarmine B 0.25 |
|
100.0ml |
|
Distilled water, cold
Dissolve, then add: |
|
1.0ml |
|
Glacial acetic acid
Filter
or |
|
1.0g |
|
Azocarmine G |
|
100.0ml |
|
Distilled water
Boil, cool, then add: |
|
1.0ml |
|
Glacial acetic acid
Filter |
|
|
|
|
| Solution B |
|
1.0g |
|
Aniline blue, W.S |
|
0.5g |
|
Orange G |
|
8.0ml |
|
Glacial acetic acid |
|
100.0ml |
|
Distilled water
Filter |
|
|
|
|
| Aniline Alcohol |
|
0.1ml |
|
Aniline |
|
100.0ml |
|
Ethyl alcohol, 95% |
|
|
|
|
| Acid Alcohol |
|
1.0ml |
|
Acetic acid |
|
100.0ml |
|
Ethyl alcohol, 95% |
|
|
|
|
| 5% Phosphotungstic Acid |
|
5.0g |
|
Phosphotungstic acid |
|
100.0ml |
|
Distilled water |
STAINING PROCEDURE:
- Stain in Solution A at 55ºC for 30 minutes. Then cool to 37ºC
and stain for 2 hours.
- Wash in distilled water.
- Rinse in aniline alcohol to remove plastic stain; few dips.
- Rinse in acid alcohol; few dips.
- Treat in phosphotungstic acid for 30 minutes to 3 hours.
- Rinse in distilled water.
- Stain in Solution B at 60ºC for 5-15 minutes.
- Rinse in distilled water.
- Dip in 95% ethyl alcohol for 2-3 dips.
Blow dry
Mount.
NOTE: If sections stain too long in Solution B, they begin to lose
the orange and overstain with blue.
RESULTS:
| Nuclei |
|
red |
| erythrocytes |
|
red |
| muscle |
|
orange |
| glia fibrils |
|
reddish |
| mucin |
|
blue |
| reticulum |
|
dark blue |
| glomerula stroma |
|
dark blue |
| collagen |
|
dark blue |
Warning: Some
of the chemicals used for the staining procedures given in this section
may be hazardous if misused. For this reason, read and observe all
warnings and cautions provided by the manufacturer for each chemical
before proceeding with a staining procedure.
Note: In order to prevent sections
from loosening from the slides during staining, all sections should
be heat-fixed
(60ºC to 100ºC) to the slides for a minimum of 2-5 minutes prior
to staining, preferably at the time the sections are mounted on the
slides. |
