[Histonet] frozen tissue

[Geoff McAuliffe]

Ethylene glycol is the main component of automotive anti-freeze. I would 
omit it from the cryoprotectant.

Geoff

Patsy Ruegg wrote:

>I am having trouble getting tissue to cut that are prepared as such:
>
>"Animals were heparinized through the inferior vena cava under Avertin
>anesthesia.  Hearts were removed, cannulated through the aorta, and reverse
>perfused with cardioplegia solution (physiological buffer containing high
>concentrations of KCl and EGTA) for 5 min at 2 ml/min to clear blood and
>fully relax the heart.  Perfusion was then switched to 4% paraformaldehyde
>in PBS for 5 min.  Hearts were removed from the perfusion apparatus,
>ventricles were dissected free of aorta and connective tissue, and hearts
>were stored overnight at 4C in cryoprotectant solution (0.1 M phosphate
>buffer, 30% sucrose, 30% ethylene glycol)."
>
>I took these tissues and snap froze them in liquid nitrogen in cryomolds
>filled with OCT as I usually do frozen tissue, as I tried to cut them in the
>cryostat they seemed raw as if the glycol had effected the freezing, the OCT
>surrounding the tissue was well frozen but the tissue seemed not so well
>frozen.
>
>Does anyone have experience with freezing tissues that have been place in
>ethylene glycol cryo protectant, I have not seen this before.
>
>Any advice would be appreciated.  Some of these tissues are still in 30%
>sucrose which I transferred them to to try and rinse out the glycol.
>
>Thanks,
>
>Patsy
>
> 
>
> 
>
>Patsy Ruegg, HT(ASCP)QIHC
>
>IHCtech, LLC
>
>12635 Montview Blvd. Ste.215
>
>Aurora, Colorado 80045
>
>Phone: 720-859-4060
>
>Fax: 720-859-4110
>
>pruegg <@t> ihctech.net
>
>www.ihctech.net 
>
>www.ihcrg.org 
>
> 
>
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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583; fax: -4029 
mcauliff <@t> umdnj.edu
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