Ear'ole mounts (2nd of your 2 questions)

From:"J. A. Kiernan" <jkiernan@julian.uwo.ca>

> 2) I am doing whole mounts of hamster ears and am having trouble getting
> them to lay flat on the slide.  Any suggestions?

In the 1970s we did whole mounts of mouse ears: not the whole thing,
but the cartilage, after peeling off the skin, and the peeled-off skin.
These preps were stained by various methods, notably supravital
methylene blue. These objects, probably thinner than your hamster
specimens, could not lie flat because they were not flat. The
cartilage is elastic, even after staining, fixation, dehydration,
clearing etc and it refuses to resist having its curved form flattened.
The skin is not flat either. This may be due to persistently
retaining the form of the underlying cartilage, and/or to variation
in the thickness of the indumentum.

Having  brought a stained preparation to a clearing agent, we
placed it on top of a drop of mounting medium on a slide, poked
it with a needle (but not perforating the specimen) to tease out
air bubbles, put mounting medium on top, and then added a coverslip.
The slides were than placed on a hotplate, slightly uncomfortably
warm to the hand (?=45C), and inspected frequently while the
medium was setting. Pressure was judiciously applied if bubbles
happened. We used coverslips much bigger than the ears, so there
was plenty of space remote from the specimen that could receive
the bubbles.

In the 1980s my lab was busy with the enteric nervous system, which
has been an object of whole-mountery since before the First World
War. We did rat and mouse preps of about 2 cm squares, dissected
by a stripping technique from the layers of 2 cm lengths of the
small and large intestine. These were dried onto slides either
before or after staining. The immunohistochemistry was done before
mounting, following the general methodology of those who work with
thick frozen sections of minimally fixed rodent brain. I can't go
into the technical details here & now, but suffice it to say that
the mounting medium was used very generously.
For thick whole mounts the best mountant may be full-strength
Canada balsam, which has to be heated to be liquid enough to
use. I was lucky enough to be able to play with about 20 gm of
this in the 1970s, but it is no longer sold. (Or is it? Tell me
I'm wrong, please!) Even the more usual balsam-in-xylene medium,
which was never the best for most purposes, has become
ridiculously expensive. If I were doing thick whole-mounts now,
I think I would try a high viscosity synthetic medium such as
Cytoseal 260.

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

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