Brain dissection for immunoblot - (Jun/09/2011 )

how do you dissect brain tissue for immunoblot analyzis, especially if you like to analyze phsophoproteins? I do not mean LASER dissection but with scalpel and binocular... any suggestion is welcome!

Inmost sun on Thu Jun 9 10:20:24 2011 said:

Get dry ice and crush it until its a fine powder. as soon as the brain is removed, place it (ventral side up) on a microscope slide, and cover with the powder to freeze the outermost layer to get it stiff enough to make coronal sections with a razor blade. cut off the parts you dont need so you end up with a coronal section containing your region of interest. lay that down on the chilled slide. You can do this all on wet ice, but ideally you use dry ice and work quickly. If it gets too warm the brain goes to mush. We use micro forceps (#5) from Fine science tools http://www.finescience.com/Special-Pages/Products.aspx?ProductId=335&CategoryId=29
one in each hand. Pluck out your sample (Paxinos brain atlas helps to learn morphology) and put it in a tube that is already in dry ice. I touch the sample to dry ice as soon as its out as well, to freeze it. This works fairly well for phopho proteins on western blot.