[Histonet] Rodent processing and artifactual spaces

Jackie O'Connor b427297 <@t> aol.com
Thu Aug 2 12:35:34 CDT 2012


I think the processing times are fine, sorry Rene'.   They are pretty similar to my processing schedules for both species.  It could be more of a fixation artefact as you suspect.  Tissues will begin to degrade waiting for formalin to get to them, and although formalin most tissues at about 3mm per hour - whole organs tend to slow penetration down somewhat. The low volume of fixative to tissue only compounds the problem.  
Jackie O'



-----Original Message-----
From: Rene J Buesa <rjbuesa <@t> yahoo.com>
To: Katherine Murphy <katherine.o.murphy <@t> gmail.com>; histonet <histonet <@t> lists.utsouthwestern.edu>
Sent: Thu, Aug 2, 2012 12:03 pm
Subject: Re: [Histonet] Rodent processing and artifactual spaces


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

_______________________________
rom: Katherine Murphy <katherine.o.murphy <@t> gmail.com>
o: histonet <@t> lists.utsouthwestern.edu 
ent: Thursday, August 2, 2012 11:07 AM
ubject: [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,
I, spleen) and have been seeing a separation artifact in the tissues
esp. kidney, brain, and heart) on the slides. I believe this is what
as described as artifactual spaces between individual cells or cell
hrinkage, as described by Carson (1997) under Fixation and
rocessing. The processing schedule that has produced this is: 50%
eagent Alcohol (RA), 70% RA, 80% RA, 95% RA x 2, 100% RA x 2, xylene
 2, and Paraplast Plus @ 60°C with vacuum, at 40 min per station for
at and 25 min per station for mouse. Preceding processing is a 20’
ap water rinse. Tissues have been in formalin for weeks in most cases
ut not always collected ideally (low formalin to tissue ratio, small
ars used for large tissues, tissues submitted whole without slices or
cores, etc)--we don't have much control over this part. Does anyone
now what causes the separation artifact is and how it can be
orrected?
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