Protocol Online logo
Top : New Forum Archives (2009-): : SDS-PAGE and Western Blotting

SDS-PAGE for cell culture medium - sample preparation - (Jun/07/2012 )

Hi, everyone! I recently encounter a problem in SDS-PAGE for mammallian cell culture medium.

I was doing a transient expression of a target protein (~16kDa) in CHO cell using DMEM+ 10% FBS medium. I collected the culture medium and load 10μl onto 4%-12% Tris-Glycine gel together with Novex Protein Sharp Prestained Standard. During the electrophoresis, I observed that the last three bands of ladder (15kDa, 10kDa and 3 kDa) were packed in a 3mm clear zone across the whole gel. And after staining, there is a big bulk of albumin.

What can i do to remove this bulk of albumin and concentrate my samples without using any kits?


There are chemical depletion methods out there that I believe rely on selective precipitation, but you run the risk of losing non-albumin proteins as well. If your protein of interest is low in abundance, kits are really the way to go.