Hello,
I would go for His-tag purification and I was wondering which company would be the best to order Ni-TA column? Do you have a preferred one or all of them would be the same?
Thanks in advance!
Judit
Dear Judith
i think that each scientist as one different prefered resins and probably there is not one best of the others in absolute terms, it is sa balance beetween costs, flow, aspecific binding, total binding capability and so on..
Generally i'd like to use Ni-NTA FF from Qiagen for purification of small sample volumes using gravity flow coloumns (i packed small coloumns from a volume of resin that can change from 200ul to 2ml). I like this resin because it is a real fast flow and it works well in gravity mode with out request high speed sample centrifugation after lysis.
When i have yo use prepacked coloumns generally i like the GE HIstrap FF crude (5ml each) because using it, you do not need to load in it surnatants from ultracentrifugation or filtered at 0.22uM and therefore are most simple to use.
Take in consideration that you can perform IMAC also using different metals from Nickel, as copper, zinc and cobalt.
I like also use zinc that is less toxic, but it work quite well. The only disadvantage is that zinc did not colour the coloumn and therefore you do not see the colour that confirm to you the binding of the metal but you have to trust it.
i wish that this information can be usefull to you in some way.
let me know if you would like more details.
good luck
Manuele
Hi Judit,
to use the old adage, "No-one was ever fired for buying GE columns". Consistent quality and reliable performance, even when recycled after many uses, although they tend to be more expensive.
I have also used Ni / Co / Zn IMAC resins from ABT in Spain and had good results with these - their prices are also quite competitive. website is : https://www.abtbeads.es/
Good luck, Robin
One of the most critical points is actually which metal ion you use. Zn often gives the weakest binding and copper the strongest. This wil especially influence the binding of contaminating host cell proteins. If you are using a hexahis tag, zinc will certainly result in complete binding. Ni is the most common, but here you need to be careful with the elution. Many other proteins will also bind and then you must control the specificity of the process by eluting stepwise.
Good luck
Leif
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