[Histonet] Rodent processing and artifactual spaces

[Rene J Buesa]

Rat and mice tissues do not have too much fat and are susceptible to dryness after processing specially when using xylene as "clearing" agent.
I also think that your times are too long.
The best processing protocol for rodent tissues is a sequence of 2-propanol → propanol+mineral oil → mineral oil → paraffin.
It has produced great results in those labs that are using it.
If you are interested I can send you under separate cover the procedure.
René J.


________________________________
From: Katherine Murphy <katherine.o.murphy <@t> gmail.com>
To: histonet <@t> lists.utsouthwestern.edu 
Sent: Thursday, August 2, 2012 11:07 AM
Subject: [Histonet] Rodent processing and artifactual spaces

Hello Histo Gurus, I have a question for you:

I process rat and mouse tissues (kidney, liver, lung, brain, pancreas,
GI, spleen) and have been seeing a separation artifact in the tissues
(esp. kidney, brain, and heart) on the slides. I believe this is what
was described as artifactual spaces between individual cells or cell
shrinkage, as described by Carson (1997) under Fixation and
Processing. The processing schedule that has produced this is: 50%
Reagent Alcohol (RA), 70% RA, 80% RA, 95% RA x 2, 100% RA x 2, xylene
x 2, and Paraplast Plus @ 60°C with vacuum, at 40 min per station for
rat and 25 min per station for mouse. Preceding processing is a 20’
tap water rinse. Tissues have been in formalin for weeks in most cases
but not always collected ideally (low formalin to tissue ratio, small
jars used for large tissues, tissues submitted whole without slices or
scores, etc)--we don't have much control over this part. Does anyone
know what causes the separation artifact is and how it can be
corrected?

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