-----Original Message-----
From: Rowani [mailto:rowani.mohdrawi@student.adelaide.edu.au]
Sent: Mon 9/15/2003 12:08 AM
To: histonet@pathology.swmed.edu
Cc:
Subject: [Histonet] Mice embryonic brain - EM Poor tissue quality - need help

Hi
My electron microscopy sections of brain of mice foetuses (18.5 dpc)  are
difficult to interpret because of  disrupted and lysed cells. The connective
tissue surrounding the cells  appears disintegrated with empty
spaces/vacuoles like artifact ?.  The brain was processed according to the
following protocols: brain was dissected and immersed in ice cold fixative
(glutaryldehyde 2.5%, paraformaldehyde 4%, sucrose 2% in PBS pH 7.2) for 1-2
hrs to firm it up before cutting them to smaller fragments (2 cumm). After
an overnight fixation, re-fixed in osmium tetroxide, washed in PBS,
dehydrated through a series of ethanol (70%, 90%, 95% and 100%) and embedded
in araldyte resin.  The brain of adult mice processed using similiar
protocol was excellent.  COuld someone suggest ways to improve the tissue
quality ?

TQ

Rowani




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