osmiumtetroxide enhancement of immunocytochem.

Noone at uclink.berkeley.edu Noone at uclink.berkeley.edu
Thu Aug 17 11:06:16 EST 1995

>>I have done immuno against parvalbumin in brain tissue and staining is present
>>but weak.  I've read that one can use osmium tetooxide to enhance visualisation
>>of the precipitate from the DAB reaction.  However, I need specific
>>instructions about how to use the osmium to do this.  If you can help me out,
>>post in the
>>news group or email me at kopl   that is kopello at eden.rutgers.edu.
>hi Michael,   I can't provide the method but I thought you could do with
>some OH & S stuff. Osmium tetroxide is mega- dangerous. It is both a strong
>poison &  mutagenic.

I don't recommend using Osmium tetroxide to enhance DAB staining, unless you want to investigate your tissue under the electron microscope. Osmium tetroxide is used for post-fixation for electron microscopy (without it the tissue would shrink a lot during dehydration; it is hard to see membranes even after contrasting with uranyl acetate and lead citrate, etc.) and it happens to enhance DAB also.

If you need to see your staining only under light microscope, try other methods instead. For example, Nickel Ammonium Sulfate is very good for DAB (it was popular in the late eighties - I have not followed the literature ever since). As far as I know, you need the usual carefulness because you work with DAB-stained sections, but Nickel is not as dangerous as Osmium. Silver can also enhance DAB staining (although Janssen distributes silver solution to enhance their colloidal gold), but it is harder to control. You have to know, however, that if you don't see nice staining with your DAB alone under a good light microscope (e.g. nice staining is when you can follow axons and see synaptic boutons on them) then metal enhancement won't save you. Metal enhancement is generally used to make a nicer contrast; the sections are then easier to photograph.

I don't have the exact recipes with me here (sources, concentrations, time, temperature, light level in the room while staining with DAB and enhancing with metals, etc.) but you could start with the following reference: Gaykema RP; Gaal G; Traber J; Hersh LB; Luiten PG.
     The basal forebrain cholinergic system: efferent and afferent connectivity
     and long-term effects of lesions.
   Acta Psychiatrica Scandinavica. Supplementum, 1991, 366:14-26.

Good luck,

Gyongyi Gaal

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