[Histonet] removing cryoprotectant from fixed tissue

From:Karl Garsha


I'm trying to help someone who has some formalin-fixed tissue sections 
that have been cryoprotected in a 30% ethylene glycol/20% glycerol 
solution; we'd like to process the tissue for laser microdissection for 
purposes of RNA extraction. The difficulty is that cryoprotectant tends 
to absorb the laser wavelengths and heat the tissue during the cutting 
process, causing damage and decreasing the liklihood of successful RNA 
extraction (the likelihood of which has already been reduced by the 
fixation process).

I would imagine that there must be a way to de-cryoprotect through 
successive washes in a miscible solvent, I'm not sure if this would 
cause even greater complications with RNA work, or if fairly gentle 
standard protocols exist so I'm appealing to the collective knowledge of 
this list:) Has anyone experienced a need to remove cryoprotectant from 
tissue (brain) sections and are there perhaps methods to accomplish this 
that might be somewhat amenable to later extraction of RNA from select 
features in the tissue sections?

Thanks in advance for any advice or helpful hints.

Sincere Regards,

Karl Garsha
Research Microscopy Specialist
US-Southwest Region
Leica Microsystems-Life Sciences Research

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