[Histonet] Mice heads in Paraffin

[Thomas, Nancy]

Mote,
For the E17-19, we found that they need more time in paraffin.  We have 2 hours in each of the dehydration  and clearing steps and two hours in each of the 4 paraffin baths.  This is 8 hours in paraffin, but for this larger size, that is what we found to work.

Nancy Thomas 
Stowers Institute for Medical Research
Kansas City, MO

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of mote uncle
Sent: Thursday, February 16, 2012 8:28 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Mice heads in Paraffin

Hello

This is my first post to this site. I have encountered a problem with embedding of mice heads in Paraffin (Gurr). Initially with Embryonic day 13-14, the protocol was working properly and the sections were also nice but with Embryonic day 17-19 I could see even after several hours of ethanol wash and xylene treatment the mice heads were soft but still I embedded the samples in paraffin. When I removed the paraffin blocks from the mold I could see the mice portion to be white, some flecks of wax coming out of surface and the surface of wax had some cracks. I could not rule out what was causing the problem.

Protocol I used for E 17-19

After Fixing in EDTA (2 weeks) and putting the mice heads in running water for 48 hours,

50% Ethanol 3 hours

70% Ethanol 3 hours

95% Ethanol 3 hours

100% Ethanol 3 hours

100% Ethanol 3 hours

Xylene 2-4 hrs (till clearing of samples was observed)

Paraffin 30 mins

Paraffin 30 mins

Paraffin 30 mins

Paraffin 30 mins
Could anybody share me your protocol for Embryonic day 17-19 and Post natal days.
Regards
Mote
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