[Histonet] diastase digestion

[Tony Henwood]

Yep,

You can still spit if you like but there is an alternative. See: V-M.
Mangan, V. Farago, M. Kelly, and A. F. Henwood (2002) " An Amylase
Reagent with a Long Shelf Life for the Removal of Glycogen from Tissue
Sections" J Histotechnol. 25(3): 153-4.

Apart from blowing my lab's own trumpet, this procedure digests glycogen
in 10 minutes at room temp and the solution lasts several months at 4oC:

Amylase Reagent
Warning: Harmful, contains azide - see MSDS
Alpha Amylase from Bacillus Subtilis (Fluka Cat No 10070,)	1g
Oxoid PBS Tablets (Cat No BR14a)				1 tablet
Distilled water							100ml
Sodium Azide							0.1g
This solution, once prepared is stored at 4oC when not in use. A
recycled antibody dropper bottle (often used in commercial
immunoperoxidase kits) is useful for storage and application.

Regards

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: 612 9845 3306
Fax: 612 9845 3318




-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Wiese,
Jason VHAROS
Sent: Thursday, 11 January 2007 9:22 AM
To: Akemi Allison-Tacha; Histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] diastase digestion


Dinosaur?  I graduated from my HT program in 2000.  I am but a puppy to
most of you big dogs.  However, In the past 6 years of working as a
bench tech, I have found no better digestion method then spitting on the
slide.  As far as "the green stuff after lunch being too much for the
youngsters"... try an autopsy with necrotic bowel after lunch, or why
not before lunch.  

We work in pathology and some of us think spit is gross?  Maybe now is a
time to re-evaluate your career choice while you are still young.  I was
in diapers when Bonnie Proctor taught you this most excellent method.  I
used it today.  Is there a problem with this as far as CAP is concerned?
I have it in my protocol.  Even though we all have a different enzyme
make up, I have yet to see different end results from different people's
spit.  So, as long as I am not breaking some cardinal rule I say...
haccc toooowie... :)


Jason Wiese, BS, HT(ASCP)
Histology/Cytology
Roseburg OR 
541-440-1000 Ext. 44751






-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Akemi
Allison-Tacha
Sent: Wednesday, January 10, 2007 1:21 PM
To: Histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] diastase digestion

Hi,
  I was one of those dinosaurs. Years ago, I used to use the old spit
method too.  Nothing like natural enzyme digestion.  I was taught that
method  by Bonnie Proctor from Providence Hospital, Portland OR in 1976.
She was Navy trained.  It worked great, but the green stuff after lunch
got to be a little too much for the younger histo tech's.  Plus, it's
not exactly a set standard protocol, since everyone has different enzyme
make-up. You also had to rinse it off for a long period of time because
it dried out on the parameter.
   
  Diastase malt didn't seem to work very well after years of
inconsistent results.   Our lab switched to amylase digestion.  We got
more consistent results.  Check with sigma-aldrich.com 
   
  Good Luck,
   
  Akemi Allison-Tacha BS, HT(ASCP)HTL
  President
  Phoenix Lab Consulting & Staffing
  Specializing in Histology, IHC & TMA
  Madison, WI
  Cell: (925)788-0900
  E-mail: akemiat3377 <@t> yahoo.com

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