Re: [Histonet] Excessively wrinkled sections

From:"Gudrun Lang"

It would help to put the sections first in a room temperature waterbath.
(small glass). There you can flatten it carefully with a brush. Mount it on
a slide and let it slowly into the warm water.
It is a easy method for people that learn cutting paraffinsections.
We let the slides for a few minutes on the border of the waterbath to dry.
The temperature of the waterbath is 40 degrees.

greetings from Austria
Gudrun Lang

----- Original Message -----
From: "Barlow, Gillian" 
To: "'Histonet'" 
Sent: Thursday, September 25, 2003 8:00 PM
Subject: [Histonet] Excessively wrinkled sections

> Dear Hisonetters
> First, many thanks to those of you who replied to help me with my cardiac
> fixations - I now have nice, well-fixed tissues.  However, one problem
> remains - the sections are very wrinkled and do not flatten out well on
> waterbath, even when left for long periods of time.  I store my blocks at
> room temperature, but put them on ice for at least two hours before
> sectioning.  The waterbath is at 40 degrees.  A colleague suggested
> increasing the waterbath temperature, but the sections are for IHC and I
> dont want to damage my antigen.
> Many thanks
> Gillian
> PS  For anyone out there with questions on cardiac fixation and embedding,
> this is the protocol I have now found to work well and wanted to share:
> Fixation:
> Hearts were isolated from anaesthetized animals, cut in half and fixed
> overnight in 10% buffered formalin at room temperature.  The following
> hearts were transferred to 70% ethanol and stored at 4 degrees prior to
> embedding.
> Embedding:
> 80% EtOH, 45 mins room temperature
> 95% EtOH, 45 mins room temperature, two changes
> 100% EtOH, 45 mins room temperature, two changes
> Xylene, 45 mins room temperature, two changes
> Paraffin, 45 mins, 60 degrees under vacuum, two changes
> Embed immediately
> Gillian Barlow, PhD
> Postdoctoral Fellow
> Laboratory of Julie Korenberg, PhD, MD
> Cedars-Sinai Medical Center
> Davis Bldg, Lab 2007
> 110 George Burns Rd
> Los Angeles, CA 90048
> Phone: (310) 423 7650
> Fax: (310) 423 0302

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