You can see a protein with silver staining but not with Coomassie staining? That is really weird.

How long do you stain the gel in Coomassie? Do you use self-made Coomassie stain (is your recipe/solution ok?) or do you buy it? Do you want to stain several small proteins which togheter are 30 µg or do you have one small protein?

I stain SDS-Pages with self-made Coomassie solution  (0,25 % Coomassie Brilliant Blue R-250) for one hour and destain them over night. Then I can see small proteins (11 kDa ) with an amount of 1 µg - but only if the whole protein amount is in localisated in one band.

Check your Coomassie solution and stain the gels longer.

Good luck and greetings from Germany,

Katharina

Looks like most of your protein is proteolytically degraded. The small portion of the intact protein that is left is not enough to be detected by Coomassie but enough to be revealed by Silver Staining. When your protein is analyzed in total serum it is protected from proteolytic degradation by many protease inhibitors present in the serum. When you partially purified your protein you might remove those protease inhibitors but not proteases themselves (especially if some purification steps are performed at room t°. So, I would recommend to include protease inhibitor cocktail at each step of your purification beginning from the total serum.

Thank you for your answer. I will take into acount the protease inhibitor cocktail for the new assays. 

Also I can made a new Coomassie solution with 0.25% of Coomassie Brilliant Blue R-250 becouse I have used 0.1%. Thanks Katharina