[Histonet] RE: Help needed with mouse brain

[D'Andreas Williams]

Greetings, 

 

We are currently trying to optimize our processing protocols for mouse
tissue on a VIP 6 processor. Of particular concern to us is mouse brain
processing. For the mouse brain we have noticed "fraying" of the edge of
the brain. The "fraying" looks a lot like  what we see when tissue is
over processed. The brains cut as if they are overprocessed; they are
brittle and crunch when they hit the blade. At times they roll up and
create a hole in the ribbon as our histologists cuts them. Along with
the "fraying" edges we noticed that the middle of the brain looks
underprocessed. When the section is placed on the waterbath the section
wrinkles greatly and spreads considerably, and if left longer than a few
seconds explodes. We have tried several protocols, and we see this
effect. The brains we are using are cut at about 4 mm thick, and they
are sectioned at 5 microns at a temperature of 42 degrees. Below are
links to 3 images of a brain. Image 1 shows the edge "fraying". Image 2
show the wrinkles in the tissue. The tissue was left as long as possible
on the waterbath to remove wrinkles and avoid breaking apart. Image 3
shows the brain breaking apart before the wrinkles have had a chance to
work their way out. Does anyone know what is causing this issue? Also, I
have put together a document of all the protocols we have ran with
images of the results that I can send to you. Thank you in advance.

 

https://dl.dropbox.com/u/25427596/Histology/IMG_1682.JPG

 

https://dl.dropbox.com/u/25427596/Histology/Wrinkles.jpg

 

https://dl.dropbox.com/u/25427596/Histology/Center.jpg

 

 

 

With Kindest Regards, 

 

D'Andreas Williams

Research Associate

Cell Biology & Bioimaging Core 

Pennington Biomedical Research Center

dandreas.williams <@t> pbrc.edu