[Histonet] Tape Transfer Methods For Cryosectioned Brains

[Patpxs]

Hi Heather,

I can see why you’re having trouble. 30 micron sections are inherently unstable.  Like paraffin, the thicker a section is the more difficult it is to cut.  Plus since your practice samples are old they tend to be more brittle. 

Try cutting at 5 microns and see what happens. Remember to let the samples warm up in the cryostat if they have been stored at -70 or -80 degrees.  If they are too cold they are brittle too.  

I have not tried the tape method.  We just place the sections directly on the slide.  

Paula

Sent from my iPhone

> On Apr 15, 2021, at 11:24 AM, Heather Deziel <hdeziel at cnscro.com> wrote:
> 
> 
> Hi Paula,
> 
> I am cutting at -24, but have tried going as warm as -18.  I am currently learning with 30uM sections with the ultimate goal of moving towards 5 or 10uM.  Our lab standard has been collecting into millonigs solution and doing free floating IHC.  We generally have no issue with this technique, but do lose some of the peripheral damaged tissue near the infarct in our stroke brains.  We're trying to work up painting the samples directly onto slides and skipping free floating staining to get a better end product.
> 
> My current samples are very old, they were collected into 4%PFA in 2017 and cryoprotected by freezing in OCT cryomatrix in a little dish floating on LN2 in 2019. They've been stored at -80 since then.  Usually we process the brains within a few weeks of collecting them so this particular tissue isn't our ideal situation. We're using old tissue to practice technique, so the current samples aren't going to be used for any actual analysis.  When we heard about the tape method of collecting we were very curious as the the opinion other labs have about it.  Have you tried it?
> 
> Thanks for the answer!
> 
> Heather
> 
> Heather Deziel, MSc.
> Laboratory Technician, CNS|CRO
> 550 University Ave, Charlottetown, PE C1A 4P3
> 
>                   (a subsidiary of Neurodyn Life Sciences Inc.)
>  
> 
> 
>> On 2021-04-15 10:29, Patpxs wrote:
>> 
>> Good Morning Heather,
>> 
>> I have some  questions about how you cut frozen brains.   
>> 
>> What temperature are you cutting at?
>> How thick are your sections?
>> How are your samples frozen?  Flash freezing, slow freezing, iso-pentane in LN2? 
>> 
>> Your answers may provide clues to help you get better cryosections. 
>> 
>> Paula
>> 
>> Sent from my iPhone
>> 
>>> On Apr 15, 2021, at 5:39 AM, Heather Deziel via Histonet <histonet at lists.utsouthwestern.edu> wrote:
>>> 
>>> Hello Histonet, 
>>> 
>>> I'm looking into working up a tape transfer method of collecting
>>> cryosections of brain while preserving infarct to be used in IHC.  I
>>> find that when I try and section heavily damaged regions of the brain
>>> the tissue tears and and I lose it.  Has anyone got any recommendations
>>> about the the Section-lab transfer tape (Kawamoto method), using the
>>> circuit plating tape recommended here
>>> (https://www.future-science.com/doi/full/10.2144/btn-2018-0021) or the
>>> cryojane system from Leica?
>>> 
>>> Thank you, 
>>> 
>>> Heather
>>> 
>>> Heather Deziel, MSc.
>>> 
>>> Laboratory Technician, CNS|CRO 550 University Ave, Charlottetown, PE C1A
>>> 4P3 
>>> 
>>>                  (a subsidiary of Neurodyn Life Sciences Inc.)
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