[Histonet] Re-embed agar/FFPE tissue in plastic?

[Morken, Timothy]

Kate, we regularly re-embed paraffin-embedded tissue in epoxy resin for electron microscopy. We are using very small tissue - kidney, liver, 1mm core biopsies. 

For that we run thru xylene 4 x 30 min at 60C, 100% ethanol 3 x 10 min, 95, 70, 50% ethanol 5 min each, then to distilled water, then buffer. Then it goes on to processing for resin embedding. The wax is gone and will not interfere with the plastic. The agar will not be affected or affect plastic embedding. We use agar to embed cilia for EM with good results. 


You might want to do some test tissue to get the timing right before trying on your samples. 


Tim Morken
Supervisor, Electron Microscopy/Neuromuscular Special Studies
Department of Pathology
UC San Francisco Medical Center


-----Original Message-----
From: Kate Davoli via Histonet [mailto:histonet at lists.utsouthwestern.edu] 
Sent: Wednesday, May 29, 2019 8:12 AM
To: Histonet Usegroup
Subject: [Histonet] Re-embed agar/FFPE tissue in plastic?

I have a bunch of precious FFPE samples that were embedded in agar prior to
being embedded in wax (so as to orient the tissue easily).  The client now
wants these samples to be taken backwards out of wax and processed for
semithin plastic GMA (JB-4) sectioning.

Is that possible? Will the tissue having previously been embedded in wax
cause problems for the reagent infiltration or the plastic curing?  Will it
interfere with the catalyst?  I know that taking tissue back through
xylenes and alcohols is *supposed *to be able to remove all the wax, but
does it really?

I need the agar support surrounding the sample to STAY on the sample so
that it can stand upright during the plastic curing ... has anybody tried
to do double embedding with agar and then JB-4?  Does that work?

Please help!
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