Home User resources Answers to frequent questions Sample preparation questions I want to use silver staining. What protocol should I use?
I want to use silver staining. What protocol should I use? PDF Print E-mail
Attached is a paper describing how to stain a gel with silver stain so it will have a better chance of working with MS. Omitting the gluteraldehyde is the critical step. Silver does other things to a protein as well so in some cases, even using this procedure will not assure success, but we have had successful id's of protein from some silver stained samples if people follow this protocol.


Anal Chem. 1996 Mar 1;68(5):850-8.
Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels.
Shevchenko A, Wilm M, Vorm O, Mann M.
European Molecular Biology laboratory (EMBL), Heidelberg, Germany.

Again, we usually discourage users from submitting silver stained gels for the following reasons:
  1. Traditional silver stain protocols oxidize amino acid side chains and introduce glutaraldehyde cross-links, interfering with sequence determination.  There are kits may reduce amino acid oxidation, but the other reason is, 
  2. Silver staining overestimates the amount of protein that's actually present in your sample.   We still need picomoles (or 100s of femptomoles at the very least) to identify your protein.  If you want to identify post-translational modifications (especially phosphorylation) we will need 10X ~ 100X as much material.
Please also look over our protocols section and the discussion of sample preparation variables affecting the success of a Protein identification run.  Please  This e-mail address is being protected from spambots. You need JavaScript enabled to view it  before running or staining your gels if you have any questions.
 

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