[Histonet] Re: Histonet Digest, Vol 45, Issue 21

[daydawning <@t> wideopenwest.com]

Caroline,

I think you require a vibrating blade microtome.  It is designed to cut 
fresh tissue while submerged in cooled buffer. You attach the fresh brain to 
a disk with (believe it or not) crazy glue and the blade slowly slices 
through the tissue and it floats off into the buffer.  It is more gentle on 
the tissue than freezing and sectioning in a cryostat.  I'm sure you could 
find someone in your facillity that has one.  Check out microm's website.  
the model is the 650V

Dawn Truscott, HT(ASCP)
Biocare Medical
Personal Email



> 
> Message: 19
> Date: Wed, 15 Aug 2007 00:46:41 -0400
> From: Caroline Bass <cbass <@t> bidmc.harvard.edu>
> Subject: [Histonet] quick and easy sectioning of fresh brain
> To: Histonet <histonet <@t> lists.utsouthwestern.edu>
> Message-ID: <756D65C0-7DC5-4302-A4FC-BAF3AFA8CEB2 <@t> bidmc.harvard.edu>
> Content-Type: text/plain; charset=US-ASCII; delsp=yes; format=flowed
> 
> Hey Folks,
> 
> So I want to have my cake and eat it too.  I have injected a rat 
> with  a virus that will express EGFP.  Ideally I'd like to collect 
> the  fresh brain, cut off thick floating sections, enough to get a 
> feel  for the spread of the virus, and possible punch out the EGFP 
> area to  get mRNA and/or protein for qPCR and western blot.  Are 
> there any  suggestions for doing this?  I have access to cryostats,
>  sliding  microtomes, and brain molds.  Cutting fresh brain is 
> difficult, so  I'd like to freeze it to section on the sliding 
> microtome, collect a  floating section, and visualize under a 
> fluorescent dissecting  microscope.  Are there any issues with doing 
> this?  How should I  freeze the brain, and how will this affect mRNA 
> and protein levels?   How difficult are unfixed floating sections to 
> deal with?  Finally,  can I take some sections and fix by immersion 
> in PFA so that I can  store for longer periods of time.
> 
> Any suggestions would be appreciated.  I've had very good luck  
> working with perfused/fixed tissue, and would like to continue with  
> it, but it doesn't seem possible with the mRNA/protein endpoints.
> 
> Thanks,
> 
> Caroline
> 
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> End of Histonet Digest, Vol 45, Issue 21
> ****************************************
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