Palmgren 1948.
Use celloidinized 12 micron paraffin wax sections.
- Take to distilled water.
- Treat sections with acid formalin for at least 5 minutes.
- Wash in three changes of distilled water over 5 minutes.
- Place in silver solution for 15 minutes at room temperature.
- Drain the slide and place in the reducer that has been heated to 40-45°C, agitate gently and leave for 1 minute. The sections should be yellow-brown in colour.
- Rinse in 50% alcohol for 5-10 seconds then in three changes of distilled water.
- Examine microscopically, if there is insufficient impregnation repeat the procedure from the acid formalin stage. Reduce the time in the silver solution and the temperature of the reducer to 30°C.
- Tone in the gold chloride solution - 2 minutes.
- Drain and treat with intensifier - 15 seconds.
- Wash in water, fix in 5% sodium thiosulphate - 2 minutes.
- Wash again in water, dehydrate, (prolong to remove the celloidin) clear and mount.
Nerve fibres - black.
Background - grey.
Note:
The intensifier and toning stages may be omitted in which case the
results would be: axons - dark brown/black, background - golden brown.
Acid formalin
Formalin - 25ml
1% nitric acid - 0.2ml
Distilled water - 75ml
Silver solution
Silver nitrate - l5g
Potassium nitrate - l0g
5% glycine - 1ml
Distilled water - 100ml
Reducer
Pyrogallol (pyrogallic acid) - l0g
1% nitric acid - 2ml
Absolute ethanol - 550ml
Distilled water - 450ml
(allow to stand 24 hours before use)
Toning solution
Gold chloride (sodium tetrachloroaurate) - 1g
Distilled water - 200ml
Glacial acetic acid - 0.2ml
Intensifier
50% ethanol - 100ml
Aniline oil - 2 drops