Hi Chandni,

Silver staining is not a quantitative stain method.  Therefore, the more you develop it, the darker it will get.  As you are confirming, to see bands that are a lot less than the ~3-6 ng, you need to develop it longer.  This means that the background also gets darker.  I am not sure that there is a solution if you need to see all the lower amounts of proteins in the gel.

You can also try and use the stop solution (Is it the 1% acetic acid solution?) to prevent development of color after you discard the developing solution.

Good luck, Hediye.

hi 

silver staining is a very tedious process. besides the exact concentration there are lot of things to be taken care of:

the silver staining chemicals should be of very good quality. I can suggest to go for sigma mainly for the developer.

The developer solution should not be cloudy. if its cloudy, filter it,

Containers should be very clean. After detergent washing wipe them with isopropanol or ethanol. whichever available.

The water used for developer should be chilled  otherwise the bands will develop faster and there are chances of black background. If developer would be chilled then bands will develop slowly and bands will be sharp.

make sure everything you are using for the process is cleaned well.

Hope this will work.

good luck.

Hi Chandni, are you using a fixative with formaldehyde?

A high background might indicate that you are not washing the fixative from the gel well enough.

Hi Chandni,

Based on my experience,

 you should use glass plate for staining process .therefor your background will decrease.

Plastic container isn't suitable for silver staining method.!!

Good luck

Yes i have been using fixative with formalin and bands of protein which i used to clearly (2.5ug) also is not visible with clean background

Reduce your silver nitrate incubation time to 5- 10 min. Incubate this step under dark without fail. It will reduce your background.

Good luck..