Re: Osmium Tetroxide

From:Philip Oshel

A brief osmium comment: I always get the crystals, not solutions. 
Cheaper. It's easy to get the crystals into solution, though. Hold 
the *unopened* vial in warm to mildly hot tap water, and the OsO4 
will melt. Then open the vial, add water water/buffer, swirl, and 
pipette out. Repeat once or twice. Or, the opened vial with melted 
crystals can be dropped into the water or buffer into which the 
osmium is to be dissolved.
But I have to agree with the others: pay *close* attention to safety! 
Work in a fume hood, wear gloves, be careful.


Hi Anila:

Anila Syed wrote:

My question is: I am embedding corpus callosum in epoxy resin in 
order to quantify the fibres. I will stain with Toluidine Blue.I am 
going to fix in glutaraldehyde overnight at 4 deg C. But would I also 
need to use osmium tetroxide? My reasoning is that it will help to 
stabilise the lipids in the myelin sheaths and therefore give me a 
much clearer picture of what is going on.
  This is only for LM and will not go to the EM at all.

     The osmium will not only stabilize the lipids in the myelin, it 
will act as a stain giving more contrast. It will also make the 
tissue easier to infiltrate with epoxy be virtue of the action of 
osmium on membranes. The tissue will also be much easier to see in 
the resin, especially if you are working with small pieces, if it is 
osmicated. Of course, osmium does not penetrate tissue very well so 
unless your tissues are thin (2 mm or less, ideally 1 mm), the center 
of the tissue may not see the osmium. And. as someone else mentioned, 
osmium must be used under a "hood", the vapors are toxic and will 
damage the cornea of your eyes. Osmium is also very expensive due in 
part to the protective packaging required. The crystals can be 
dissolved but it does take several days.
     On the other hand, depending on the epoxy you are using, the 
toluidine blue may look "nicer" without osmium.
     You might want to look at Gerrits et al., J. Neuroscience Methods 
45:99-105, 1992. They used Sudan Black B to stain GMA-embedded 
sections of spinal cord. Beautiful results!

Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583; fax: -4029

Philip Oshel
Supervisor, BBPIC microscopy facility
Department of Animal Sciences
University of Wisconsin
1675 Observatory Drive
Madison,  WI  53706 - 1284
voice: (608) 263-4162
fax: (608) 262-5157 (dept. fax)

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