Dave,

Here's what I  can think of off hand to check:
Tissue has to be snap frozen in isopentane and artifact free.
The ammonium sulfide has to be very fresh.
The sections have to be cut evenly and without knife marks.
If you overadjust the pH, start over.

It is important that the muscle biopsy is frozen as soon as it is received.
I read an article on a study  done on muscle not  frozen immediately where
enzyme activity was not altered.  This may be true, but I'm from the old
school and I wouldn't take that chance.  We had the biopsy sent to us fresh
immediately after removal in clamps or tied to a stick.

 The lab had the stain working on precut slides that were stored in a -70
freezer.  The slides were stained as soon as they were bought to room
temperature.

This is probably one of the toughest stains to get working.  And as the
Neuro-pathologist informed us, you have to keep trying and than finally it
works for some unexplained reason.  But once it works it alway works.

Good Luck,

Rande Kline HT (ASCP)
Technical Services
EM Science




Marjorie & David Cardwell <cardwell@connexus.net.au> on 02/09/99 06:22:01
AM

To:   Histonet@Pathology.swmed.edu
cc:
Subject:  MUSCLE HISTOCHEMISTRY - problems with ATPases?




Has anyone had problems with inconsistent ATPase staining on muscles,
especially at pH4.6?

I'd be very interested to hear of people's approaches to this stain.

David Cardwell
State Neuropathology Service
Melbourne, Australia