[Histonet] freezing mouse heart tissue

[Malika Benatti]

You may found that excessive amount of saline could cause freezing artefacts, so dabe

Try to embed heart sample on cryostat chuck directly, with OCT then freeze chuck on dry ice block directly as opposed to using liquid nitrogen.

Another option, would be freezing tissue in hexane boiling tube, with Acetone/dry ice freezing mixture. 

Hope this help.

Malika 



On 26 Mar 2010, at 13:22, Merced M Leiker wrote:

> Hi David,
> 
> I wouldn't snap-freeze the tissue in the OCT blocks. For one this could cause the blocks to crack; you may have noticed this already in some of your blocks?
> 
> But you may want to try freezing the OCT block by resting the block in a tray of isopentane (2-methylbutane) that has been frozen over liquid N, as you've already thought of.
> 
> You can also search the Histonet archives on this topic as it's been discussed before here.
> 
> Regards,
> Merced
> 
> 
> --On Friday, March 26, 2010 8:26 AM +0100 David Santer <etw <@t> gmx.at> wrote:
> 
>> Hello,
>> 
>> 
>> 
>> I am currently trying to produce cryosections from mouse heart tissue. I
>> already have experience with paraffin-sections and had faced no major
>> problems. But with cryosectioning I would ask for your help. You can get
>> an idea of our current status at
>> <http://www.d-cup.at/histo/mouseheart.jpg> this link  (Hematoxylin test
>> stain, not H&E, the whole heart section looks like this) and as you might
>> guess I am not satisfied with the quality. Would you call this freezing
>> artifacts? Some people suggested that freezing with only LN2 would be not
>> quick enough and create those ice crystals.
>> 
>> 
>> 
>> Here is how we prepared the tissue:
>> 
>> After taking out the hearts from the mice, we flushed them retrogradely
>> via the aorta with cold sodium solution. Then we cut the hearts in half,
>> put them into cryomolds and covered them with OCT. Afterwards they were
>> snap-frozen in liquid nitrogen and stored at -80°C.
>> 
>> 
>> 
>> Do you have an advice or maybe a suitable protocol for me? Would you
>> recommend 2-methyl butane?
>> 
>> 
>> 
>> Thank you very much! Greetings from the sunny Vienna!
>> 
>> 
>> 
>> David
>> 
>> --
>> 
>> Mit freundlichen Grüßen
>> 
>> with kind regards
>> 
>> 
>> 
>> Dr. David Santer
>> 
>> 
>> 
>> Ludwig Boltzmann Cluster for Cardiovascular Research
>> 
>> c/o Core Unit for Biomedical Research
>> 
>> Waehringer Guertel 18-20 - Leitstelle 1Q
>> 
>> A-1090 Vienna
>> 
>> Austria
>> 
>> 
>> 
>> Website:  <http://www.cardiovascular-research.at>
>> www.cardiovascular-research.at
>> 
>> 
>> 
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>> 
> 
> 
> 
> Merced M Leiker
> Research Technician III
> Cardiovascular Medicine
> 348 Biomedical Research Building
> State University of New York at Buffalo
> 3435 Main St, Buffalo, NY 14214  USA
> leiker <@t> buffalo.edu
> 716-829-6118 (Ph)
> 716-829-2665 (Fx)
> 
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> 
> 
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