[Histonet] Phosphorylase method

[Mitchell Jean A]

Sharon: we do the phosphorylase procedure routinely on all muscle
biopsies.  It is a stain that very, very seldom have we seen an abnormal
result.  But phosphorylase is also a secondary stain to review in the
case of glycogen storage to confirm those findings.

I will forward my phosphorylase procedure to you in a separate email.
The procedure has clean results, is permanent; and can be coverslipped
with regular mounting media (I use cytoseal).   Jean 

-----Original Message-----
From: Sharon Allen [mailto:SAllen <@t> dsmanitoba.ca] 
Sent: Wednesday, March 28, 2012 11:12 AM
To: Mitchell Jean A
Cc: histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] Phosphorylase method

Thanks for the quick response. 
 I would love to take a look at your method. Our method from Dubowitz's
2nd edition has worked fine for years, but can leave residue on the
slides, especially when fresh, fades & is very messy to file and we seem
to be going back to previous bx's with more frequency. The 3rd edition
of Dubowitz & Dawson's Neuropathology Techniques do not use PVP or
glycogen, but use Dextran.  This solution is much nicer to work with,
not as thick & does not leave deposits on the slide when fresh and
claims to appear stable for several years. So, finally, after doing this
test one way "because we have always done it that way", I am trying to
clean it up a little. 
Also, do you do phosphorylase routinely, if not what is the criteria for
doing it.  At present we do it as a part of a panel for all muscle bx's
(which amounts to 80 to 110 per year). It seems this may not be
necessary.
Thanks again for your interest; I would appreciate your input.
Have a good day,
Sharon Allen

Senior Medical Technologist

Neuropathology Lab-MS435U

Health Sciences Centre

820 Sherbrook Street

Winnipeg,MB, CA 

R3A 1R9

e-mail: sallen <@t> dsmanitoba.ca
-----Original Message-----
From: Mitchell Jean A [mailto:JMitchell <@t> uwhealth.org]
Sent: Wednesday, March 28, 2012 9:07 AM
To: Sharon Allen
Subject: RE: [Histonet] Phosphorylase method

Sharon: Just curious - Does the phosphorylase method you use fade after
time?  

The method I use does use PVP and dextran (no glyceron though).  I am
not certain of why these compounds are used; may have something to do
with carbohydrate bonding but wouldn't swear by it.  I make the
incubation media up in larger batches; aliquot into smaller amounts;
freeze at -20 until ready to use.  I section my muscle onto 22 X 22
coverslips and only use 10ml aliquots of incubation media at a time. 

Your name sounds familiar; possibly from one of the muscle workshops I
have given at NSH?  Anyway - if you don't already have my procedure -
let me know and I will forward.

Jean Mitchell, BS HT (ASCP)
University of Wisconsin Hospital & Clinics Neuromuscular Laboratory
Manager 600 Highland Avenue Madison, WI  53792-5132 

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Sharon
Allen
Sent: Wednesday, March 28, 2012 8:47 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Phosphorylase method

Hi,

Does anyone have a good enzyme method for Phosphorylase done on muscle
bx's. We have been doing the method from Dubowitz, 2nd edition for years
but would like to get away from the messy PVP & coverslipping.  We are
trying cytoseal, which seems to work fine for coverslipping, but would
like to compare other methods without PVP.  Also, does anyone know the
purpose of PVP, glyceron or dextrin in this test?

Thanks for any help,

Sharon Allen

Senior Medical Technologist

Neuropathology Lab-MS435U

Health Sciences Centre

820 Sherbrook Street

Winnipeg,MB, CA 

R3A 1R9

e-mail: sallen <@t> dsmanitoba.ca