Re: [Histonet] Bielschowsky's stain

From:"Sarah Jones"

Hi Robin,
   If you have never seen this done before, it may be confusing to you.
Use concentrated ammonium hydroxide (less than one year old) and keep
adding it until the solution clears.  It takes quite a bit of ammonium
hydroxide to get it to clear, but it will.  I'm not sure of the amount
because I just use a transfer pipet and keep adding it.  Constantly
swirl the flask as you add the ammonia to the silver.  At the end it
goes fast so watch for that.  It's lots of chemistry fun and one of my
favorite solutions to make up.  

Sarah Jones HT(ASCP)
Dept. of Vet. Anatomy & Public Health
Histology Lab
Texas A&M University
College Station, TX 77843-4458
phone: 979-845-3177
fax:  979-458-3499

>>> "Robin Turcotte"  11/19/2003 6:54:58 PM >>>

Does anyone have good result with BIELSCHOWSK'S STAIN ?

I am using the technique from Bancroft as follows on brain tissue of

1. Deparaffinize and hydrate to distilled water. Wash well in distilled
2. Place sections in 50 ml of 20% silver nitrate in the dark at 37 C.
Keep this silver solution for 
stage 4.
3. Wash well in distilled water.
4. Add strong ammonia to 50 ml of silver nitrate (from stage 2) until
the initial precipitate disappears. 
Leave section in this solution for 10 minutes at 37 C.

I would to understand step 4. Should I add concentrated ammonia?

I added concentrated ammonia 1 to 2 drop at a time. Immediately I got
precipitate that did not disappear.

I repeated this step with dilute ammonia getting the same result. I
filtered the soluton and continued the
staining and obtained no color on my axons.

I guess I have a  problem with step 4. How much volume of ammonia
should be added at what concentration. Is there a critical point in this

Thank you 


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