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Silver Staining Method Compatible with Mass Spectrometry
Posted by: Vania, Northwestern University, Chicago, IL
This Silver Staining Protocol was adapted from Shevchenko A et al, Anal Chem, 1996 and allows detection of proteins on a SDS-PAGE gel with high sensitivity but at the same time allows identification of the spots/bands by MS/MS analysis.
Materials.
Fix solution: 5% acetic acid, 45% reagent alchol, 50% water
Sensitizer: , Silver and Developer reagents fresh every time.
Fix 1: 50% methanol/10% acetic acid
Fix 2: 5% methanol/1% acetic acid
Fix 3: 50% methanol
Stop: 6% acetic acid or 40 mM Na2-EDTA
SENSITIZER: 0.02% sodium thiosulfate
20 mg Na2S2O3• 5H2O + 100 ml ddH2O
Reserve a small volume of Sensitizer (2.0 mL) for Developer solution.
SILVER: 0.2% silver nitrate
200 mg AgNO3 + 100 ml ddH2O Keep at 4ºC until used
DEVELOPER: sodium carbonate/sodium thiosulfate/formaldehyde
6.0 g NaCO3 + 50 μl (37% stoc...