RE: cutting fixed "sucrose" livers

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From:"Barry, Lilith" <> (by way of histonet)
To:histonet <>
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For sectioning  wit cryostat you need to use only 15% sucrose on PB.  It is
good enough for not having large ice crystal formation as you are freezing
the tissue. It will allow you to cut at about -20 degrees C. Freezing in
liquid nitrogen though might crack your specimen. Your second option sounds
better. You can also freeze it on dry ice or in isopentane coled to
-70degrees C.
If you have more questions please contact me,
Lilith Ohannessian-Barry
National Research Council
Institute of Biological Sciences
Montreal rd Campus, M54
Ottawa, Ont. K1A 0R6

From: I.H.Straatsburg DIVG
Subject: cutting fixed "sucrose" livers
Date: Wednesday, January 13, 1999 2:16PM

To start with: thank you for the suggestion to infuse fixed tissue
with a sucrose gradient before cutting cryostat sections. Without
sucrose, the morphology of the sections was awful. Unfortunately, the
procedure I am using at the moment requires fixation of the liver
tissue prior to freezing. I would rather do without.

Here comes the problem: the liver and kidney tissue infused with 50%
sucrose in 0.1 M Phosphate buffer (until it sank) is very difficult
to cut in a cryostat (-20 degrees Celsius; 8-10 micrometers
Note: The tissue samples were not impregnated in OCT compound (I take
that is 'Tissue Tec' or something similar) before freezing, but
directly frozen either in liquid N2 or on a cryo-boost-plate in the
cryostat cabinet.

Should I cut a lower temperatures? I have got the impression, the
tissue is too soft, too sticky.
Is the impregnation with OCT compund essential?

I am very curious and thankful for any help!
Dr. I.H. Straatsburg
IWO1-155, Dept. Exp. Surg., Surgical Lab.,
AMC, Amsterdam, NL 1105 AZ
tel. +31.20.5666653

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