Hi Liuqing, The scheme you suggested (or heard) of using Ion-exchange at the end can be useful to separate protein with same net charge but different conformations.
Ciao On Fri, Nov 17, 2017 at 1:49 PM, Gianluca Cioci <gianluca.ci...@gmail.com> wrote: > 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 >>> >> >>
