[Histonet] postnatal brain sections using vibratome

[Merced Leiker]

I also recall having issues in getting antibodies to bind or epitopes to 
retrieve a year or so ago using 0.5% GA (with the PFA) in perfusing...

--On Friday, March 06, 2009 11:40 PM +0800 TF <tifei <@t> foxmail.com> wrote:

> Hi, I dont think glutaraldehyde is good for many antigens...though the
> brain does become firmer...
>
>
> 2009-03-06
>
>
>
> TF
>
>
>
> ???? Geoff McAuliffe
> ????? 2009-03-06  23:37:34
> ???? shymaa shawadfy
> ??? histonet
> ??? 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 ? 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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>>
>>
> --
> --
> **********************************************
> Geoff McAuliffe, Ph.D.
> Neuroscience and Cell Biology
> Robert Wood Johnson Medical School
> 675 Hoes Lane, Piscataway, NJ 08854
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Merced M Leiker
Research Technician II
354 Biomedical Research Building
School of Medicine and Biomedical Sciences
State University of New York at Buffalo
3435 Main St, Buffalo, NY 14214
Ph: (716) 829-6033
Fx: (716) 829-2725

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