[Histonet] Schiff mystery

[Tony Henwood]

Peggy,

I would be worried by the activated charcoal.
It could be possible that the charcoal clarifying step is removing too
much of the active schiff reagent. The students may be trying to obtain
a clear solution and mixing the charcoal one more time. I would suggest
that they stop once a yellow colour is obtained.

I had a similar problem when preparing thionine-schiffs. Activated
charcoal removed the active schiffs reagent as well as the contaminants
so for a blue schiffs I do not clarify with charcoal. The dehydrating
alcohols remove the pink-purple background staining, leaving the
positive blue staining.

Regards

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
Tel: 612 9845 3306
Fax: 612 9845 3318
the children's hospital at westmead 
Cnr Hawkesbury Road and Hainsworth Street, Westmead 
Locked Bag 4001, Westmead NSW 2145, AUSTRALIA 




-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Lee &
Peggy Wenk
Sent: Monday, 2 November 2009 9:07 PM
To: histonet <@t> lists.utsouthwestern.edu
Cc: 'Peggy Wenk'
Subject: [Histonet] Schiff mystery


Need the help of the Histonet chemistry gurus.

This has happened the last 2 times the students and I made up Schiff.
Two different set of students.

We make up the Schiff, and use it the next day, and it works correctly
(stains the glycogen in liver and neutral mucins in intestine a bright
magenta). Store in refrig. 1 week later, when we go to stain fungus with
PAS, the Schiff doesn't work. Try the same liver and small intestine
controls. Doesn't work at all. No magenta, no pink, nothing on the
slides.

When going into the water rinse after Schiff, the water turns pink with
the newly made Schiff, but 1 week later, there is no color change in the
water rinse.

When we make up the Schiff, we show the students the Schiff quality
assessment, where we put a few drops of formaldehyde in it, and the
Schiff changes to purple immediately. Works great newly made. One week
later, when we can't get the Schiff to stain, there is no color change
with formaldehyde. None.

It is the same procedure we've been using for years to make up the
Schiff. It is the same procedure our histology lab uses. Their Shiff
lasts for months. Ours last for a few days.

I think it might be one of our reagents is going bad. Histology lab has
their own reagents, the School has our own. 
- The basic fuchsin is OK, as the Kinyoun works great.
- The hydrochloric acid was taken from histology, so I'm guessing it's
OK.
- The charcoal is fairly new. Could that be the problem? Why?
- We are using sodium metabisulfite, not sodium sulfite or sulfate. At
least that's what the bottle says. Does this break down and go bad?

Need all the help we can get. This is spooky, Halloween or no Halloween.

Peggy Wenk, HTL(ASCP)SLS
Schools of Histotechnology
Beaumont Hospital
Royal Oak, MI 48073


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