Re: rat testis

From:Lorraine Gibbs

Hi Ayesha,

Make certain that you bisect the testis before fixation. I would fix at room
temperature overnight. Bouins or Zamboni's fixation are good choices for
testis. They are probably? compatible with the TUNEL assay. I would do the
graded ethanols at about 45 minutes each (after the post-fixation rinsing
step in 50% alcohol). Also, a clearing agent other than
xylene--limonene-based or other, is preferred. Paraffin times should total
about 2 hours and the paraffin should not be over-heated.

Good luck!

I do not often work with testis and hope that you will get more replies from
others. Perhaps a PubMed search on rat testis can get you more info?
www.ncbi.nlm.nih.gov/PubMed/

Lorraine Gibbs
Physiology & Biophysics
University of Washington
----- Original Message -----
From: "Ayesha Joshi" 
To: 
Sent: Tuesday, January 08, 2002 12:28 PM


>
> Hi! There...
> I am working with rat testis. I need to perform TUNEL assay on testis
> sections. The kit for TUNEL recomends fixation with 10% formalin. I have
> tried this fixative atleast 2-3 times (18-20 hrs at 4 degrees) with
> standard subsequent procedures
> but the sections I get are not good at all. The tissue is shrunk and is
> fragmented. Is there a specific requirement for processing testis? I use
> alcohol grades for a period of 1hr in each grade, followed by iso-propanol
> and then xylene as the clearing agent before paraffin embedding.
> Could people who routinely work with testis sections help me out?
>
> I am in dire need of help as my work is really stuck because of this!!!
>
> Thankyou in advance..
> Ayesha.
>
>
============================================================================
>
>
>
>   AYESHA JOSHI
>   Graduate Student
>   MRDG(CRBME)
>   Indian Institute of Science
>   Bangalore-560012
>   INDIA
>
>
===========================================================================
>
>





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