On Tue, 4 Jan 2000, Kim Burns wrote:

> I've been asked to stain spermatoza using the Heidenhain's azan
> technique. I am having trouble obtaining one of the ingredients, analine
> oil. Does analine oil have another name? Might you know where I can
> purchase analine oil?

   The liquid you need is aniline. In the AZAN method (probably the
   best of all the classical trichromes), water saturated with
   aniline is used as a weakly alkaline solution to extract most of
   the first-applied dye (azocarmine G or S) from collagen. The next
   stage of the method (PTA or PMA) completes the extraction from
   collagen but does not extract azocarmine from certain cytoplasms
   and secretory granules. The third staining step injects methyl
   blue (aniline blue WS) into collagen, including thin "reticular"
   fibers, and yellow (orange G) into erythrocytes and cytoplasms
   not already colored by azocarmine G/S.

   Heidenhain's AZAN is not a "routine" method. Every step must be
   checked under a microscope on the wet slide, and the person doing
   the checking must know what to expect to see. The reward for
   correctly carrying out the procedure is a section that shows
   nuclei, secretory granules, collagen (& basement membranes) and
   other structural features in sections that can be 10 or 12
   microns thick. Ordinary "routine" trichrome methods such as
   Mallory, Masson, Gomori or Cason demand 4 or 5 micron sections.

   If your boss requests Heidenhain's AZAN, ask him to show you
   exactly how to proceed. He should be the one to carry the can
   for correctly staining the preparations.

 John A. Kiernan,
 Department of Anatomy & Cell Biology,
 The University of Western Ontario,
 LONDON,  Canada  N6A 5C1
   E-mail: kiernan@uwo.ca