Silver Staining is a highly sensitive method for detecting proteins and nucleic acids in polyacrylamide slab gels. The silver ions interact and bind with specific protein functional groups on the protein gel. The controlling the selectivity and effectiveness of silver ion binding to proteins, as well as the successful conversion (development) of bound silver to metallic silver, requires a variety of sensitizer and enhancer reagents. Finally, Silver ions are converted to metallic silver throughout the development process, resulting in a brown-black hue.

After looking the attached stained gel, I do not find anything wrong with it. Since silver staining is very sensitive, therefore you have to be very careful while developing the gel. Yes, there is a few limitations with the protocol of the silver stain that little over development may generate high and erratic background and the extreme protein to protein variability in staining. Therefore, avoid over staining.

Thus, check all solutions as well as protocol and strictly follow it. Avoid overexposure (development) in order to get appropriate staining. For this purpose, you can reduce the exposure time in the developer.

Good luck

Hi,

I had this problem myself, I also dealt with too weak staining. If signal is to strong make the half concentration of formaldehyde and nitric acid in your solutions. It will weaken the signal.

I even managed to stain only DNA from TCL sample by reducing concentrations of formaldehyde and nitric acid, took a photograph and stained the gels properly to get complete staining (DNA, RNA and proteins).

Hello,

I think that the developing buffer concentration should be optimized. In our lab, we are using the following concentration of Na2CO3, 3 % (m/v); formaldehyde, 0,05 % (v/v). You could use different concentrations if the issue still persists.

Good luck!