Hi Liuqing, I would not recommend SEC. SEC does not give that great of a separation unless your contaminant is greatly different in size. Instead of SEC, you might want to consider hydrophobic interactions chromatography (HIC). You can add your ammonium sulfate directly to your eluted protein from your IMAC column or IEX, avoiding the dialysis step. I would also recommend trying some test kits which have a variety of columns to test and see what works best for your protein. We use these kits routinely for our clients and have good luck with HIC.
*David L. Blum, Ph.D.* Department of Biochemistry and Molecular Biology | *Director, Bioexpression & Fermentation Facility* 120 Green Street | Life Sciences Bldg A414A | Athens, GA 30602 706-542-1035 <7065421035> | b...@uga.edu | bff.uga.edu On Fri, Nov 17, 2017 at 8:44 AM, Liuqing Chen <519198...@163.com> wrote: > Hello everyone! > I have listened someone suggested that, first use affinity chromatography > (Ni-NTA), then use SEC (superdex200 increase), and finally used ion > exchange (monoQ), to purified protein, which will be used to > crystallization. > My question is why the monoQ used in the finally step, why not the SEC > used at the finally step? > > sincerely > Liuqing Chen >
