Good Morning, Behnaz.
I am curious which modification of Bielschowsky you are using. Most of
the common modifications do not require Sodium Carbonate.
See below for my protocol which is based on Hirano's modified
Bielschowsky. Please let me know if you have any additional questions.
Good Luck! jennifer
Jennifer L. Hofecker, HT(ASCP)
Vanderbilt University Medical Center
Division of Neuropathology
NSH Quality Control Committee Chair
MODIFIED BIELSCHOWSKY METHOD
1. 20% SILVER NITRATE
Silver nitrate............20 g
Distilled Water.........100 ml
Mix well and protect from light. Prepare fresh. Fill one coplin
jar and set aside.
2. AMMONIUM HYDROXIDE (CONCENTRATED)
3. REDUCED SILVER SOLUTION
Add ammonium hydroxide, drop by drop, with constant stirring to
the remaining 20%
silver nitrate solution. The solution will turn from dark brown to
black and then to clear. Filter through #1 filter paper ALWAYS
4. AMMONIA WATER
Ammonium hydroxide.........2 drops
5. DEVELOPER (PREPARE IN THIS ORDER)
Citric Acid.........0.25 g
Formaldehyde, 37%......10 ml
Nitric acid...............25 uL
Mix well after adding each item. Stable for 5 days @ room temp.
6. 5% SODIUM THIOSULFATE
Sodium Thiosulfate...............5 g
Distilled Water..................100 ml
1. Deparaffinize sections and hydrate to distilled water.
2. Place slides 20% silver nitrate at room temp. for 30 min.
Protect from light.
3. Rinse at least 6 times in millipore double distilled water. Save
solution from step
4. Wipe moisture from slides and incubate in Reduced Silver
solution. Protect from light.
5. Place slides directly in ammonia water. Save reduced silver.
(Slides can remain in ammonia water for several minutes without
harming the staining. Only 1-2 min are needed for the solution
to have desired effects.)
6. Put 3 drops of developer in 30 ml of reduced silver (do not
7. Place slides in this solution on a shaker near a microscope.
8. Watch for sections to turn brown. Rinse slides in water. Wipe
back of slide and check under microscope. Plaques and tangles
should be black. Check each slide individually. If not dark
enough, return to ammonia water for 1 minute and back in reduced
silver/developer solution. If silver starts to break down and form a
"mirror" on the coplin jar, put 3 drops of developer in another 30 ml
of reduced silver (can use solution from Step 4).
9. Wash in running water for 15 minutes. Rinse in distilled water.
10. Place in 5% sodium thiosulfate for 2 minutes.
11. Wash in running tap water for 1 minute. Rinse in distilled
12. Dehydrate through graded alcohols.
13. Clear in two changes of xylene, 3 min each.
14. Mount with synthetic resin.
Senile Plaques and neurofibrillary tangles of Alzheimer's disease -
Neurons and background - brown to yellow.
Lipofuscin - brown or black
[mailto:firstname.lastname@example.org] On Behalf Of Behnaz
Sent: Tuesday, April 01, 2008 1:55 PM
Subject: [Histonet] Alzheimer stain
Question regarding Sodium Carbonate Solution in Bielschowsky method for
Alzheimer. Can I substitute Carbonate? Or any one has a better method
for this stain? ( I do not have Sodium Carbonate, and no time to order
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