[Histonet] Problem with Schiff's reagent

[Thomas Jasper]

Hi Paul,

Some people may think this is taking the "easy" way out, but...you might
consider purchasing some commercially prepared Schiff's.  It will
(should) be consistent product (consistently produced) and the
manufacturer has the responsibility of QC'ing the product before it is
shipped and sold.

There are any number of reliable companies handling Schiff's (I would
not favor one over another).  You could type "Schiff's" into your search
engine and I'm certain you'd be able to contact a supplier.  This may be
an answer for you.

Good Luck,

Tom Jasper

Thomas Jasper HT (ASCP) BAS
Histology Supervisor 
Central Oregon Regional Pathology Services
Bend, OR 97701
541/693-2677
tjasper <@t> copc.net 

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of
pbrunko <@t> siue.edu
Sent: Saturday, February 14, 2009 9:54 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Problem with Schiff's reagent


Good Morning!

I'm a newcomer to the list, and I'll start by pointing out that I'm not
a histologist by any means.  I am a freshwater ecologist, and we're
trying to study mucus secretion behavior in freshwater snails.

Last Spring and last summer, we developed a process whereby we could
visualize snail mucus trails on glass slides using a periodic
acid-Schiff's reagent staining technique.  But now, in following the
same protocol as used last year for making our own Schiff's reagent, I
cannot get the final solution to filter out clear.

Recipe I'm using:
900 ml boiling water
10 grams basic fuchsin
25 ml concentrated HCl acid (12 M)
40 grams sodium metabisulfite
(this is essentially Sigma Aldrich's ratios, I think)

Let this sit for 24 to 72 hours, take 100 ml aliquot, add 0.75 - 1.0
gram ground activated charcoal, stir for 10 minutes, filter through
filter paper then through GF/C glass fiber filter.  Last summer I got
nice, clear (slightly
yellow) and very active Schiff's reagent.

But now I cannot seem to get the filtrate to be clear.  Even after 10
minutes exposure to ground activated charcoal and filtering, the
filtrate remains bright orange to dark red and it does not seem to stain
mucus trails very well.

All the reagents are the same as those used last summer (i.e., less than
7 months old; although the HCl is a bottle several years old from a
different lab).

Anyone have any troubleshooting suggestions?  I don't know the chemistry
very well, but the sodium metabisulfite is used for "decoloring" the
initial solution, right?  So is the metabisulfite not working for some
reason now??

Any help/suggestions would be greatly appreciated.

Cheers -

Paul

Paul E. Brunkow, PhD
Department of Biological Sciences
Southern Illinois University Edwardsville
Edwardsville, IL    USA
-------------------------------------------------
SIUE Web Mail



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