[Histonet] Sucrose cryoprection

[Anila Syed]

I used to put my tissue in gradually increasing concs of sucrose for up to 1
week *before* cutting them. But my samples were whole human brainstems, so
your perfused animals may require less time.
I used to try to get 50 micron thick sections of brainstem.

Hope that helps?


Anila
---- Original Message ----- 
From: "Pablo Sánchez Quinteiro" <psanquin <@t> lugo.usc.es>
To: <Histonet <@t> lists.utsouthwestern.edu>
Sent: Thursday, September 30, 2004 10:42 AM
Subject: Re: [Histonet] Sucrose cryoprection


> Thank for your help Gayle,
>
> I am sorry my mail was not detailed enough.
>
> Yes, I am doing frozen sections with a freezing microtome. The animals are
> perfused with Paraformaldehyde 4% and postfixed for 48 hours in the same
> fixative. Then they are transfered to Phosphate buffer.
>
> I have problems when cutting the tissue. The sections (60 microns thick)
> are apparently nice, but just after putting them into the buffer they do
> not stay flat and they fall to pieces after gentle touching with the
> paintbrush.
>
> I have seen that after sinking in sucrose the samples show a gelatin-like
> appearance. They seem
> swollen and they are transparent. Could be this related to a poor
fixation?
>
> For immunohistochemistry it is not recommended posfixation times greater
> tan 24 hours, but maybe postnatal or fetal material need longer
> postfixation time.
>
> Thanks for your interest.
>
> Pablo
>
> >We need more details on exactly what you are doing??????  and what the
> >problems are?  Are you using a cryostat?
> >
> >At 11:14 AM 9/29/2004, you wrote:
> >>Dear listers,
> >>
> >>I am cutting at the sliding microtome fetal and early postantal mice
> >>brains. Could somebody tell me if is it normal that after sucrose
> >>cryoprection the pieces of tissue show a gelatin-like appearance? I have
> >>lot of problems cutting this tissue.
> >>
> >>Thanks in advance
> >>
> >>Pablo
> >>
> >>
> >>
> >>
> >>_______________________________________________
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> >>Histonet <@t> lists.utsouthwestern.edu
> >>http://lists.utsouthwestern.edu/mailman/listinfo/histonet
> >
> >Gayle Callis
> >MT,HT,HTL(ASCP)
> >Research Histopathology Supervisor
> >Veterinary Molecular Biology
> >Montana State University - Bozeman
> >PO Box 173610
> >Bozeman MT 59717-3610
> >406 994-6367 (lab with voice mail)
> >406 994-4303 (FAX)
> >
> >
> >
>
>
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