[Histonet] Sections falling off slides

[mkundu <@t> purdue.edu]

Hi ,

I am trying to save the thin section of  biomaterial from falling off in 
antigen retival steamer technique.

I am using positive charged superfrost slides and I am doing regular 
microtomy with paraffin blocks .
Is there any thing else I can do in addition to prevent from falling?
The sections are very small.

Any suggestions to improve are welcome.


Thank You
MK
----- Original Message ----- 
From: "Brian Dias" <brian.dias <@t> mail.utexas.edu>
To: <histonet <@t> lists.utsouthwestern.edu>
Sent: Saturday, November 06, 2004 10:13 PM
Subject: [Histonet] Sections falling off slides


> hi
>   i'm trying to optimize my immunohistochemistry protocol on slides and am 
> runnning into a few problems.
> 1. i'd like to know if anyone has any strong opinions (either 
> positive/negative) about conducting such protocols on slides v/s 
> free-floating. i'd like to be able to do this on slides because mounting 
> my sections after performing a free-floating technique is virutally 
> impossible due to the fragile nature and small size of of my sections.
>
> 2. i run into the problem of my sectiions falling off the slide whilst 
> doing my ICC
>
>    the brain is dissected form the skull post-perfusion with saline and 4% 
> paraformaldehyde
>    it is then stored in 4% paraformaldehyde for 3 days at 4 C
>    then transferred to 20%sucrose/1X PBS and stored at 4 C for 3 days 
> before sectioning on the cryostat at a 30-50um thickness and mounted on 
> Fisher Probe-On Superfrost Plus slides. slides are stored at -80C after 
> they've dried (approx. 20 mins).
>    just before (the night before/2 hours before) performing the ICC, i 
> remove the slides from the freezer and allow to air-dry.
>    immerse slides in 4% paraformaldehyde for 10 mins at RT in a dish
>    1X PBS wash at RT in a dish
>    then normal ICC steps by overlaying solution on slides and here's where 
> sections come peeling off.
>
> i'd appreciate any input on the matter
>
> thanks a ton
> regards
> brian
>
>
>
>
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>                                                     Brian George Dias
>
>    Graduate Student (Crews lab)            Office Phone #: 512-475-6738
>    The University of Texas at Austin        E-mail: 
> brian.dias <@t> mail.utexas.edu
>    Institute for Neuroscience                  Website: 
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