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Various questions about Silver stain

Various questions about Silver stain - Protocols and Methods Forum

Various questions about Silver stain - Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.


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Old 03-07-2004, 02:06 PM
Fabius
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Default Various questions about Silver stain



I use Silver Stain for RNa and DNA in Polyacrylamide Gels and in Agarose
gel.
In the Polyacrylamide gels i use the "Merril" methods (Science, vol.
211,1427-1428, 27 march 1981) that was write for Proteins(in bief: fix the
gel, rins the gel, soak in a solution of potassium dichromate, wash the
gel, soak in a solution of silver nitrate, rins in developer solution
until the desired inensity, stop solution) but work very well also for
nucleic acids.
Instead to fix the gel in the 50 % methanol and 12% acetic acid I use 40%
ethanol- 10% acetic acid for 20'. After i use 9% ethanol-1% acetic acid to
conserve the gel at 4C before Stains, but realy I don't know wich are the
best solution for fix and conserve the gel before Stain (Someone can help
me?) .
Besides I use Ethanol instead of methanol only becouse is more cheap and
less toxic, but i don't know why Merril instead suggest methanol (Why
methanol?).
Often the stain is more yellow than black, why? Do it hit with ethanol?

In the agarose gel i try to use the methods of Zalazar et all*(to see the
note) with little result for the background.
The methods is this:
submerge the gel in 18 mM Silver nitrate- 10% acetic acid for 10' or more;
to rins in wather more times;
to develop in 3% NaOH-o,5 mL/L of 37% formaldehyde- 2mg/L sodium thisulfate;
stop solution (10%acetic acid).
Someone use this method or other methods for obtain a good Silver Stain of
DNA in Agarose gels (the problem is the background)?
I've read too that is possible to drying the gels with cellophane sheets,
but it's not clear if i can do this (withaut new machine ), how i can do
this and if drying the gel can be useful to improve the stain.

Sorry for my english, i hope to be clear,
thanks .
--
Fabio
FabioDB#deleteme#@katamail.com
-----------------------------------------------
Note:

*Modified Silver Staining for RNA and DNA in Agarose Gels. Analytical
Biochemistry 291, 299-300 (2001) and
A Procedure of Silver Staining for Nucleic Acids in Agarose Gels without
Pretreatment or Drying Steps, Analytical Biochemistry 252, 15-18 (1997)

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