Re: [Histonet] Keeping frozen mouse aorta on slides

From:Rene J Buesa

  The collagen nature of aortas make them more difficult to adhere.
  I think that you will need a "mechanical barrier" to restrain the sections. Try doing the whole procedure with the slides flat and the sections covered by a very fine plastic mesh that will hold the sections down by its weight. The mesh will allow all the reagents to act.
  The only problem will be when lifting the mesh that the sections may stick to it.
  The procedure could also be done through a piece of filter paper, but it is more likely to peel the sections than with the mesh.
  Just a thought, you may try 1 or 2 sections to see what happens.
  René J.

"Randolph-Habecker, Julie"  wrote:

I need your suggestions! I am working with an investigator who wants my
lab to stain unfixed, OCT embedded mouse aortas. Unfortunately, they
have already cut all of the tissue so we do not have a choice of slides.
They were cut at 6 microns on Star frost slides and stored at -80C for a
few months. When we received them, we took a few slides out, let them
air dry at room temperature for 24 hours, put them in a slides rack,
soaked them in room temperature PBS for 5 minutes and fixed them with
10% NBF for 10 minutes. These steps are done without agitation. This
procedure is compatible with our staining protocol and normally works
fine when we use Super frost or Gold Plus slides from Erie. However, in
this case, the tissue is floating off the slides in both the PBS and the
NBF stage. I have also tried to do these steps with the slides flat and
get the same results. 

Any suggestion on ways to "glue" already cut frozen sections to a
charged slide in ways that are compatible with x-gal and IHC staining?

Thanks in advance for your help!!!!


Julie Randolph-Habecker, Ph.D.
Staff Scientist - Director
Experimental Histopathology Shared Resource
Fred Hutchinson Cancer Research Center
1100 Fairview Ave, N. M5-A803
Seattle WA 98109-1024

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