Muscle freezing: Temperature

<< Previous Message | Next Message >>
From:Tim Morken <> (by way of histonet)
To:histonet <>
Content-Type:text/plain; charset="us-ascii"

The best way to get a consistent freezing  temperature of the isopentane
used in freezing muscle biopsies is to use a digital thermometer. We
used one from Fluka. The thermometer has a metal probe rod which can
also be used to stir the isopentane. We let the temperature go down to
neg 160 deg C and then plunged in the muscle sample for 25 seconds. It
is important, in my experience, to have the liquid nitrogen level be at
or above the level of the isopentane in the beaker in order to get the
best temperature drop.

The muscle sample (human) was prepared by sectioning off a piece of a
biopsy and orienting it so we would get a cross section. A wood square
about 25 mm square and 5 mm thick was spread with a mix of 10 percent
gum tragacanth which was made into a pyramid shape. The muscle was put
on the tip of the pyramid and pushed into the gum a little bit to hold
it. We then held it upside down until the temp of the isopentane was
just right. This insured a good orientation. We used a pair of big
forceps to hold the block.

After freezing we put it in a zip lock bag in the cryostat (if it was to
be sectioned that day) or the minus 70 deg C freezer (if it was to be
done later). Never had a single bad prep this way.

Tim Morken, B.S., EMT(MSA), HTL(ASCP)
Infectious Disease Pathology
Centers for Disease Control
1600 Clifton Rd.
Atlanta, GA 30333


FAX:  (404)639-3043

----Original Message Follows----
Date: Fri, 23 Oct 1998 10:09:35 -0700
From: "Garcia, Vicki {USA~Palo Alto}" <>
Subject: RE: Freezing Muscle
To: 'Margaret Gondo' <>,

Hi Margaret-
I just started freezing mouse soleus muscle and have tried everything!
procedure that works best for me is attaching the muscle to the chuck
gum tragacanth and then freezing it in isopentane cooled by liquid
I suspend a stainless steel beaker above the container of liquid
nitrogen so
that the very bottom of the beaker is submerged in the liquid nitrogen.
is critical that your isopentane is cold enough.  The isopentane will
to turn white at the bottom of the container and will begin to get
At this point it is cold enough to freeze muscle.

Vicki Garcia
Roche Bioscience
Tissue Repair
Palo Alto, Ca.
> -----Original Message-----
> From:	Margaret Gondo []
> Sent:	Friday, October 23, 1998 9:39 AM
> To:
> Subject:	Freezing Muscle
> Hi Kids!
> I'm going to be doing some work with muscle(frozen sections)  pretty
> I've always heard horror stories about freeze artifact and things like
> that. I just wanted to know if there is any good reference out there
> so I can read up on muscle techniques.
> Thanks,
> Margaret

Get Your Private, Free Email at

<< Previous Message | Next Message >>