Dear All The ion exchange has the great advantage ,over other tecniques, to concentrate the protein.
Histrap+sec is a big classic in protein purification but times it is worth considering other schemes. Why doing a SEC? It is for removing aggregates or a contaminant ? It the latter case probably an ion exchange or an Hic would do a better job. For fast changing of the buffer desalting with a big column is the way to go. Good luck Gia Il 17 Nov 2017 18:57, "David Blum" <dlb...@gmail.com> ha scritto: > 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 >> > >
