[Histonet] Mouse staining for ATP

[Nails, Felton]

 I am looking for a reliable method for staining mouse muscle with ATP'ase pH 4.3, 4.6 and 9.4.
Any help will be greatly appreciated.

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Tom McNemar
Sent: Wednesday, February 09, 2011 9:05 AM
To: 'Erin Sarricks'; histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] PAS with Diastase Digestion

Spit  for 5 minutes....  old school....


Tom McNemar, HT(ASCP)
Histology Co-ordinator
Licking Memorial Health Systems
(740) 348-4163
(740) 348-4166
tmcnemar <@t> lmhealth.org
www.LMHealth.org


-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Erin Sarricks
Sent: Wednesday, February 09, 2011 7:38 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] PAS with Diastase Digestion

Hi Histonet-



I recently ran a PAS/D stain and had some issues with it.  Both "with" and "without" slides came out looking the same so I'm guessing my digestion step didn't work!  I used a Malt Diastase solution (0.5g to 500mL water) for my digestion.  This is the procedure I used:



1.    Deparaffinize and hydrate to water.

2.    Place the sections labeled “with” in diastase solution preheated to
37˚C for 1 hour.  Hold the sections labeled “without” in distilled water.

3.    Wash in running water for 5 minutes

4.    Place all section (with and without) in 0.5% periodic acid solution
for 5 minutes

5.    Wash in 3 changes of distilled water

6.    Place in Schiff reagent for 15 minutes

7.    Wash in lukewarm tap water for 5 minutes (immediately sections turn
dark pink color).

8.    Counterstain in Mayer’s Hematoxylin for 3 minutes.

9.    Wash in tap water for 10 minutes

10.   Dehydrate starting with 95% ETOH, clear, and coverslip.



I am wondering if my solution possibly got too warm in the oven and hindered the enzyme activity, or is it possible I left it in too long?  Any tips would be much appreciated!  Oh, and I have about 300 slides to stain, so spitting on them is my last last last resort!  Haha!  Thanks in advance for all your help!







Erin Sarricks, HT (ASCP)

Histology Laboratory Technician

USAMRICD Comparative Pathology Branch

Office: Bldg E-3081 Room 178

E-mail: erin.p.sarricks <@t> us.army.mil
_______________________________________________
Histonet mailing list
Histonet <@t> lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet

This e-mail, including attachments, is intended for the sole use of the individual and/or entity to whom it is addressed, and contains information from Licking Memorial Health Systems which is confidential or privileged. If you are not the intended recipient, nor authorized to receive for the intended recipient, be aware that any disclosure, copying, distribution or use of the contents of this e-mail and attachments is prohibited. If you have received this in error, please advise the sender by reply e-mail and delete the message immediately. You may also contact the LMH Process Improvement Center at 740-348-4641. E-mail transmissions cannot be guaranteed to be secure or error-free as information could be intercepted, corrupted, lost, destroyed, arrive late or incomplete, or contain viruses. The sender therefore does not accept liability for any errors or omissions in the contents of this message, which arise as a result of e-mail transmission. Thank you.

------------------------------------------------------------
CONFIDENTIALITY NOTICE:
 The information in this e-mail may be confidential and/or
 privileged.  If you are not the intended recipient or an
 authorized representative of the intended recipient, you
 are hereby notified that any review, dissemination, or
 copying of this e-mail and its attachments, if any, or
 the information contained herein is prohibited.  If you
 have received this e-mail in error, please immediately
 notify the sender by return e-mail and delete this e-mail
 from your computer system.  Thank you.
============================================================