Re: Re: [Histonet] postnatal brain sections using vibratome

[TF]

Hi, I dont think glutaraldehyde is good for many antigens...though the brain does become firmer...


2009-03-06 



TF 



发件人： Geoff McAuliffe 
发送时间： 2009-03-06  23:37:34 
收件人： shymaa shawadfy 
抄送： histonet 
主题： Re: [Histonet] postnatal brain sections using vibratome 
 
If you put a little glutaraldehyde  (0.25%) in the fixaitve you perfuse 
with but not in the post fix solution the brain will be noticably firmer 
and your antigen may survive.
Good luck!
Geoff
shymaa shawadfy wrote:
> Dear all
>
> I am trying to use vibratome 50 祄 thick sections for immunofluorescence
> using Postnatal day 0 brains. The problem is that brains are very soft and
> are usually destroyed upon handling and the agarose is separated form the
> brain.
>
> My used protocol was: perfusion with 4 % PFA for 3 min, followed by several
> hours to overnight post-fixation. Then embedding brains in 2 % low melting
> agarose and cutting the block on vibratome using low speed.
>
>  
>
> I am thinking to add an overnight  20 % sucrose incubation step following
> the post-fixation step. Then embed in agarose and continue the normal
> protocol.  May be sucrose will increase the elasticity of the tissue. 
>
>  
>
> So what do you think ?
>
>  
>
>  
>
> Thanks a lot
>
> shymaa
>
>  
>
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-- 
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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
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mcauliff <@t> umdnj.edu
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