Naphthoic Acid Hydrazide
for Aldehydes
Materials
Veronal acetate
| Material | Amount | |
|---|---|---|
| Sodium acetate | 1.943 | g |
| Sodium barbiturate | 2.943 | g |
| Distilled water | to 100 | mL |
Veronal buffer pH 7.4
| Material | Amount | |
|---|---|---|
| Veronal acetate solution | 5 | mL |
| M/10 hydrochloric acid | 5 | mL |
| Distilled water | 60 | mL |
NAH
| Material | Amount | |
|---|---|---|
| 2-hydroxy-3-naphthoic acid hydrazide | 0.1 | g |
| Ethanol, 100% | 95 | mL |
| Acetic acid, glacial | 5 | mL |
Fast blue B
| Material | Amount | |
|---|---|---|
| Fast blue B salt | 0.1 | g |
| Veronal acetate buffer (pH 7.4) | 100 | mL |
Tissue Sample
5 µ paraffin sections of neutral buffered formalin fixed tissue are suitable. Many other fixatives are satisfactory. Fixatives containing strong acids should be avoided if the intent is to demonstrate aldehydes generated from acid hydrolysis of DNA, as acids in some fixatives may hydrolyse the tissue during fixation (picric acid in Bouin’s formal-picric-acetic mixture, for example).
Protocol
- Bring sections to water via xylene and ethanol.
- Oxidise or hydrolyse to generate aldehydes.
- Rinse briefly with distilled water.
- Rinse briefly with 50% ethanol.
- Place into NAH solution at room temperature for 3-6 hours.
- Rinse with three changes of 50% ethanol, about 10 minutes each.
- Wash well with water.
- Place into pre-cooled fast blue B solution for 1-3 minutes at 0°C.
- Wash well with water.
- Optionally, counterstain appropriately.
- Dehydrate with ethanol, clear with xylene, and mount with a resinous medium.
Expected Results
- Aldehyde sites – blue to purple
- Background – as counterstained
Notes
- Sodium barbiturate is also known as veronal.
- Procedures for producing aldehydes include those for acid hydrolysis of DNA, periodic acid and chromic acid oxidation of carbohydrates. In those procedures, begin at step 3, above, where those other methods specify placing into Schiff’s reagent.
Safety Note
Prior to handling any chemical, consult the Safety Data Sheet (SDS) for proper handling and safety precautions.
References
- Culling, C F A, Allison, R T, Barr, W T, (1985).
Cellular pathology technique., Ed. 4., p. 187
Butterworths, London, England.
