[Histonet] Murine brain perfusion
scoop <@t> mail.nih.gov
scoop <@t> mail.nih.gov
Fri Mar 30 10:35:45 CDT 2007
I do mouse perfusion using a 24 gauge tubing adaptor (no sharp ends)
in the left ventricle and cut the right atrium open (you need
somewhere for the perfusion fluid to exit the sytem). But I have
always wanted to try cannulating the aorta or vena cava - what size
catheter or tubing do people use for that? Also, is it possible to
cannulate the aorta through the left ventricle?
>It has been years, but here goes. We used a 2%
>paraform+1% glut mixture in a 0.1M buffer (a
>Sorensen's phosphate was our choice) at about pH 7.2.
>The setup was fairly simple: a drip bottle was filled
>with warm (actually RT) fixative and elevated to about
>6ft. A second bottle held saline for doing a flush
>and the 2 bottles were connected via a simple 2-way
>valve. A 26ga needle was inserted into the ascending
>aorta and the abdominal aorta was cut to allow
>outflow. The flush is run until clear (probably no
>more than 30-60sec) at which point the valve is
>switched to the fixative. Fixation is adequate after
>about 5min (max). The brain is removed and placed in
>fresh fixative for at least 1hr. (This technique
>should actually give whole body fixation.) Coronal
>sections were taken of the brain and either processed
>for electron microscopy, into paraffin for routine
>H&E, placed on a vibratome and 50um sections obtained
>for tracer localization (evaluating blood-brain
>barrier integrity), etc. Probably the diciest part of
>the technique is inserting the cannula correctly into
>the ascending aorta and clamping it in place--we used
>small vascular clamps for this.
>Another life, many years ago.
>Roger Moretz, Ph.D.
>Dept. of Toxicology
>Boehringer Ingelheim Pharmaceuticals, Inc.
>--- Jackie M O'Connor <Jackie.O'Connor <@t> abbott.com>
>> How do you do it? We would like to perfuse whole
>> mouse brains with
>> fixative (what kind?) in order to make 2mm coronal
>> sections to visually
>> examine the brain for therapeutic changes as soon
>> after harvest as
>> possible. Would someone be willing to walk me
>> through their protocol?
>> I don't have any experience with this at all - and
>> they're looking to me
>> for answers.
>> Happy Friday - Jackie
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