[Histonet] Cryoprotection

[Alan Bright]

Dear Pierre,

Brain should be sectioned at -8 to -12ºC  and if possible have the knife & anti-roll plate around -20ºC. You are way too cold @ -30ºC Liver & Kidney should be no lower than -18ºC lower and you will just get dust.

I hope this helps if any more info is required please get back to me. Happy sectioning III

Best Regards

Alan Bright

Bright Instrument Co.Ltd.
St Margaret's Way
Huntingdon
Cambridgeshire
PE29 6EU
England

Tel No:+44 (0)1480 454528
Fax No:+44 (0)1480 456031
Email: abright <@t> brightinstruments.com
Web Site: www.brightinstruments.com
Skype User ID: dazzle0


-----Original Message-----
From: zwiegers [mailto:zwiegers <@t> interchange.ubc.ca] 
Sent: 10 June 2006 01:22
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Cryoprotection


Good day all,

Recently, we have sacrificed and perfused a batch of mice, and due to inattentiveness, some tissues (brain, spinal cord, liver, and kidney) were protected in 3% sucrose (instead of 30%). These were frozen and stored at -80 C for over a week. As soon as we started slicing the brain(s), we noticed that the tissue was shattering. At first we tried to reduce the temperature by keeping the tissue at - 30 C overnight. The tissue was still shattering when we tried to slice it. Now, since we used expired OCT, we assumed that this could have caused the problem. We then tried to use new OCT on a batch of mice that were not frozen yet (using 3% sucrose)and the results were the same.

Fortunately, we have not lost all of our tissue and they will just be transferred to a 30% sucrose solution. Now, the question is, what can we do with the frozen tissue? Is it at all possible to thaw the OCT  and transfer the organs to a 30% sucrose soltution with minimal damage to tissue integrity? 

Thank you so much in advance

Sara Sarband
Pierre Zwiegers


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