On Fri, 16 Jul 1999, Ian Montgomery wrote:

>                                 ....  Now the problem, I have to use
> the same technique on free floating sheets of smooth muscle. Any hints and
> tips on how I can prevent the tissue shrinking from a full, but delicate
> sheet, into a wee wizzened lump.

  If your smooth muscle sheets are, by any chance, layers of gut
  wall from small animals, the following may be of interest.

   Heinicke, E.A. & Kiernan, J.A. (1990). An immunohistochemical study
     of the myenteric plexus of the colon in the rat and mouse. Journal
     of Anatomy 170: 51-62.
   Kiernan, J.A. & Heinicke, E.A. (1989). Sennosides do not kill
     myenteric neurons in the colon of the rat or mouse.  Neuroscience
     30: 837-842.
   Heinicke, E.A. & Kiernan, J.A. (1990). Resistance of myenteric
     neurons in the rat's colon to depletion by
     1,8-dihydroxyanthraquinone. Journal of Pharmacy and Pharmacology
     42: 123-125.

  In these investigations we handled the layers of longitudinal
  + circular muscle in the same way as thick free-floating frozen
  sections. We did immunohistochemical tests for several 
  neurotransmitter-related antigens.

  Antibodies to tyrosine hydroxylase stood out conspicuously in
  that they always worked well, whoever the supplier. We mostly used
  a PAP method (2 or 3 different suppliers), and occasionally ABC
  with Vector's kit. The end reaction was always a simple DAB reaction
  for peroxidase, without any kind of enhancement.

  I have a feeling in my descending colon that something in the TH
  molecule tickles up antibodies that love to be immunohistochemically
  detected. The only down side is that TH is common to all three
  catecholamine neurotransmitters. In the gut, noradrenaline is the
  most probable one around blood vessels, sphincteric muscle, or 
  neurons in ganglia of the myenteric plexus.

 John A. Kiernan,
 Department of Anatomy & Cell Biology,
 The University of Western Ontario,
 LONDON,  Canada  N6A 5C1